Chiasma-induction and tyrosine metabolism in locusts

The production of a gregarization pheromone has been postulated in locusts, with effects on melanization of the hopper cuticle and increased chiasma frequency during meiosis in the adult on crowding or gregarization. Lack of chiasma-inducing effect of the pheromone on albino strains is correlated with the absence or deficiency of some of the products of the metabolic pathways of tyrosine. Some of these products, commercially obtainable, are the amino acids phenylalanine and tyrosine leading to both the melanization and sclerotization pathways; dopamine formed from dopa in the lastnamed pathway; three products of dopamine i.e. protocatechuic acid, noradrenaline and adrenaline. The injection of solutions of these metabolites into the haemolymph of solitary hoppers has shown that only dopa to some extent but noradrenaline to a large extent are effective in raising chiasma frequency in solitarised individuals of normal-coloured strains of Locusta, while in two albino strains, which differ genetically, the injection of dopa, dopamine, protocatechuic acid and noradrenaline proved effective; phenylalanine was effective in only one of these albino strains, while adrenaline was effective in neither. The chiasma-inducing effect of noradrenaline, common to the three strains, is accompanied in the normal-coloured strain by a greater retention of dark coloration during solitarization and by some attainment of the crowded type of morphometric ratios which is a third physical criterion of gregarization. The genetic blocks to the physical criteria of gregaria in the albino strains lie at the immediate level of dopa production or previous to this reaction; it may be construed that such a block in the solitaria of normal-coloured strains also lies at this early level, in this case being induced by too low a pheromone concentration.     

The gregarisation pheromone of locusts

Swarming locusts show three physical criteria, i.e. the phase changes of melanisation of the nymphal stages or hoppers, of the proportions of certain body parts (morphometric ratios), and increased genetic recombination (meiotic chiasma frequencies) in the adult. The control of these changes, initiated by aggregation into swarms, i.e. gregarisation, seems to be vested in a pheromone which is produced by all hoppers in both the solitaria and gregaria phases, also by hoppers of the albino strain. Such a pheromone can be extracted from the locust room air and from the locust, these extracts showing high activity in bioassays, primarily in increased chiasma frequencies but also in hopper colour. The extract in risella oil is more efficient than that in petroleum ether and can be distilled to yield an active distillate. The pheromone is secreted in the faeces of hoppers but not of adults. There is evidence in faeces bioassays that all three physical criteria are affected; the pheromone may be called locustone. It is manufactured or secreted in a specific section of the alimentary canal, i.e. the crop. Reception is not through the antennae but through the stigmata. Preliminary chemical analysis of a risella oil air extract distilled into various other solvents showed the presence of a relatively simple saturated aliphatic chain with a carbonyl function, perhaps a ketone or an ester.     

Genetic and environmental factors affecting chiasma formation in locusts

By means of crosses with the African Migratory locust it has been demonstrated that various strains carry different segregations of genes which quantitatively condition the formation of chiasmata during meiosis. These genes occur in polygenic series, and in one low frequency strain the polygenes show an epistatic effect for low chiasma frequency, while another low frequency strain seems to have attained this property through gamma radiation. In reciprocal crosses between the lastnamed low and a medium frequency strain there appears to be a maternal effect, with heterosis and epistasis respectively in the reciprocal crosses. In one strain two lines were selected for plasticity and non-plasticity of hopper colour change in small populations, and this selection has resulted in multiple changes in that the two lines now diverge in the three main physical criteria of phase transformation, viz. hopper colour, adult morphometric ratios and chiasma frequencies; one line is now solitarious for these characteristics, the other more gregarious. The genotypes selected out could be related to the ability to utilize the gregarization pheromone.A 10° rise in temperature above 22° C has the effect of increasing chiasma frequencies significantly, but increase in the carbon dioxide content of the atmosphere has no effect. Male and female hoppers produce equivalent amounts of the chiasma-inducing pheromone, while solitarised hoppers do not lose the ability to produce this pheromone. The crucial stage for the action of this pheromone on chromosomes is during the fifth nymphal instar. The phase status of parents appears to affect chiasma frequencies in the offspring, more specifically that of the mother, so that there is further evidence of maternal effect on chiasma frequency.     

A locust pheromone: Locustol

For some time a gregarizaiton pheromone has been postulated, and demonstrated, to account for the changes which take place in solitary locusts when they are crowded, i.e. for phase transformation. The airborne substance which is given off by locust faeces has been extracted and submitted to ultraviolet, infrared, mass, and nuclear magnetic resonance spectroscopy. The results indicated a derivative of guaiacol, which is a degradation product of the metabolism of the lignin of plants. This airborne substance is 2-methoxy-5-ethylphenol and has been synthesized and bioassayed on solitary phase locust hoppers. It proved positive in action on the quantitative phase traits such as increased chiasma frequencies during meiosis, melanization of hopper integument, changed adult morphometric ratios, and the behaviour of hoppers: this pheromone has been named locustol.     

Some effects of amputation of the antennae on pigmentation, growth and development in the locust, Schistocerca gregaria

ABSTRACT. Removal of the antennae from Schistocerca gregaria early in the third or fourth instar resulted in fifth instar nymphs with a green haemolymph and cuticle. Antennectomized adults had a lower somatic dry weight and a smaller fat body than operated control insects. In addition, their rate of sexual development increased and some of their morphometric parameters were altered. Implantation of corpora allata into fourth instar nymphs induced a green coloration of the haemolymph after a few days. It is suggested that antennectomy of Schistocerca nymphs resulted in an increased effectiveness of the corpus allatum hormone with its concomitant effects on nymphal pigmentation, growth and development, and adult morphometries. It is further suggested that antennectomy of crowded locusts may mimic, for them, the uncrowded condition, by reducing sensory input.     

Effects of juvenile hormone treatment on phase changes and pheromone production in the desert locust,Schistocerca gregaria (Forskal) (Orthoptera: Acrididae)

The roles of juvenile hormone III (JH III) on phase changes and pheromone production were examined in laboratory-reared gregarious desert locust, Schistocerca gregaria (Forskal). The hormone was applied to 5th instar nymphs and newly emerged adult locusts. Generally, the 5th instar nymphs exhibited a higher sensitivity to hormone treatments than the adults. Hormone applications inhibited pheromone production (as measured by the amounts of phenylacetonitrile released). In addition, JH III had a significant effect on the external colouration and absorbance ratios of the haemolymph pigments. It is concluded that the effects of exogenous JH III on gregarious locusts represent a shift towards the solitarious phase.     

Density-dependent physiological phase in a non-migratory grasshopperAiolopus thalassinus

Aiolopus thalassinus thalassinus (Fabricius) (Orthoptera: Acrididae) is a non-migratory grasshopper of widespread geographical distribution, also endemic in the Tel-Arad region of the Northern Negev of Israel, where it is liable to sporadically damage agricultural crops. Periodic sampling in uncultivated ‘batha’ and agricultural fields, conducted during 1990/1991, indicate thatA. thalassinus populations exhibit seasonal fluctuations in density. Local spatial and temporal distribution, within this region, are dependent on food availability. Field observations and laboratory studies suggest that the local population ofA. thalassinus exhibits genetic heterochromy unaffected by density. The duration of nymphal developmental is 34 days for nymphs reared in isolation, whereas crowded nymphs complete their development within only 21 days. No striking density related changes in gross morphometric features are evident, but, in adults from isolated culture, an increased abundance ofsensilla coeloconica, presumably involved in olfactory chemoreception, may be functionally related to enhanced (23.6-fold higher) activity of nymphs reared under crowded conditions. This higher level of activity is correlated to higher levels of energy reserves in the haemolymph — lipids and carbohydrates — and to increased respiration. Finally, the individual food consumption of nymphs from crowded culture is almost 5-fold higher than that of isolated nymphs and may increase the potential for crop damage. These results are similar to those obtained with the migratory locustLocusta migratoria and support the hypothesis that non-migratory grasshoppers exhibit some density-dependent physiological characteristics of locusts, but do not exhibit overt chromatic or morphometric phase characteristics.     

A role for ecdysteroids in the phase polymorphism of the desert locust

Abstract. Locusts show density-dependent continuous phase polymorphism; they appear in two forms or phases, gregarious and solitary, and there is a continuous range of intermediates between the extreme phases. Although earlier studies showed that there are no major phase-dependent differences in the titres of ecdysteroid in the haemolymph of desert locusts, Schistocerca gregaria , recent studies showed some minor differences in the timing of the main peak of ecdysteroids. In crowded penultimate- and last-instar hoppers, peak titres were lower but longer-lasting, whereas in isolated hoppers they were higher but of shorter duration. The major component of the haemolymph peak of ecdysteroid was 20-hydroxyecdysone in both isolated and crowded hoppers, but differences were found in the relative amounts of two minor components (makisterone A-like compound and highly polar products). In S. gregaria adults, the regression of the prothoracic glands was irregular and subject to high individual variations, but phase-dependent differences in the rate of regression were significant, and the adult glands did not produce physiologically significant amounts of ecdysteroids. Peak titres of ecdysteroid in the haemolymph were higher in isolated than in crowded adults. Similar to larvae, adults of the solitary phase contain more ecdysone in the haemolymph than those of the gregarious phase. Moreover, the phase characteristic titres of ecdysteroid in the adult stage can be shifted from one phase to another phase in response to appropriate changes in density. In contrast, the maximum amount of ecdysteroids in both ovaries and eggs was significantly higher in the gregarious than in the solitary phase. The amounts, and to some extent the types of ecdysteroids, were the only difference between ovaries and eggs from solitary and gregarious locusts. In addition, in newly hatched larvae, the amount of ecdysteroid was more than five times higher in gregarious than in solitary phase.     

Phase transformation and chiasma formation in locusts

The chiasmata in the eight longest of the eleven autosomes in spermatocytes of three species of locusts are greatly increased from the low basic number characteristic of the grasshopper-like or solitaria phase to higher numbers in the swarming or gregaria phase. Swarming in the field gives a greater impetus to the increase in chiasma frequency than crowding in cages. Although there seems to be some maternal effect acting on chiasma formation, the main correlation between population density and chiasma frequency seems to be effected during the last, or last two, instars of nymphal life. The effective factor is postulated to be present in the atmosphere, i.e. the air circulating in the locust breeding room and that circulating around a swarm in the field.In the species Locusía migratoria migratorioides three genetic factors have been found to control chiasma frequency. The first is a dominant factor reducing the frequency in one strain, the second is a possible polygenic complex reducing chiasma frequency in another strain selected for plasticity in hopper colour change, and the third is the recessive albino mutation which maintains chiasma frequency at the same level in crowded and solitary individuals.     

The trans-generational phase accumulation in the desert locust: Morphometric changes and extra molting

To understand the underlying trans-generational phase accumulation, a classical morphometric characteristic, the F/C ratio (F, hind femur length; C, maximum head width), of adult desert locusts (Schistocerca gregaria) was monitored over eight consecutive generations. Adult F/C ratios, which are larger in solitarious locusts than in gregarious ones, were negatively correlated to the darkness of body color at hatching. Two successive generations were required for a complete shift from the gregarious (crowd-reared) to the solitarious (isolated-reared) phase and vice versa in the laboratory. That is (1) female adults needed to be exposed to crowded (or isolated) conditions so that their hatchlings would become large (or small) and dark (or green) in color, and (2) the hatchlings then needed to be exposed to crowded (or isolated) conditions for their entire nymphal stage. Solitarious locusts exhibited extra molting that influenced the F/C ratio in the adult stage, but did not exert significant influences on the trans-generational changes in this trait because the incidence was low. The incidence of extra molting was negatively correlated with nymphal survival rates. The morphometric trans-generational changes may be explained without assuming any accumulating internal factor.     

Phase-related morphological changes induced by [His7]-corazonin in two species of locusts, Schistocerca gregaria and Locusta migratoria (Orthoptera: Acrididae)

The effects of a neurohormone, [His(7)]-corazonin, on phase-related morphological traits (F/C and E/F ratios; F = length of the hind femur, C = maximum width of the head; E = length of fore wing) were re-examined in the desert locust, Schistocerca gregaria Forsk?l. The F/C ratio was significantly different between adults with five and six nymphal instars, respectively, indicating that they need to be analysed separately. Injections of the synthesized peptide (1 nmol) into individually-reared (solitary) nymphs at the second and third instars caused a shift in classical morphometric ratio towards the value typical for crowded (gregarious) individuals in both sexes. The E/F ratio, which is smaller in solitary locusts than in gregarious ones, was also influenced significantly by injections of [His(7)]-corazonin into individually-reared locusts. The effect of [His(7)]-corazonin on E/F ratios was shown more clearly when the nymphs were injected at a higher dose (2 nmol) at the beginning of the third instar. Single injections of the peptide into individually-reared nymphs at different instars revealed that the earlier the injection the larger the 'gregarizing' effects of the peptide on F/C and E/F ratios. The same tendency was also detected in Locusta migratoria Linnaeus. These results supported the hypothesis that [His(7)]-corazonin plays an important role in the control of phase polymorphism in locusts.     

Novel cross-stage solitarising effect of gregarious-phase adult desert locust (Schistocerca gregaria (Forskål)) pheromone on hoppers

Previous studies had demonstrated stage differentiation in the cohesion (aggregation) pheromone systems of the desert locust, Schistocerca gregaria. In laboratory arena, the nymphal and adult stages responded aggregatively to their own pheromone, but dispersed evenly within the arena in the presence of the other. In the present study, we explored the effects of longer-term contact of field gregarious hopper bands and laboratory crowd-reared nymphs with the major constituent of the adult pheromone. During the first few days, hoppers in treated bands became relatively hyperactive. Over the next few days, their movements became random and they stopped marching as coherent groups, they started to roost for longer periods on vegetations, and they fragmented into smaller and smaller groupings and individuals. When attacked by birds, they demonstrated subdued levels of collective defensive behaviour compared to normal hoppers, and there were clear signs of increased predation and cannibalism at the roosting sites. In cage experiments, crowd-reared nymphs treated with the pheromone component became hyperactive, showed abnormal diel patterns and reduced feeding on plants but increased cannibalism. Our observations show that the major adult pheromone constituent has a solitarising effect on gregarious hoppers. The mechanism underlying this effect and the potential of the agent in desert locust control are discussed.     

An investigation of the possible presence of an aggregation pheromone in the milkweed bugs, Oncopeltus fasciatus and Lygaeus kalmii

ABSTRACT. The presence of an aggregation pheromone was studied in two species of milkweed bugs. The nymphs, young adults and old adults of Oncopeltus fasciatus were tested for their aggregation in response to acetone extracts of themselves and each other. The O. fasciatus nymphs were also tested for their response to odour from live O. fasciatus nymphs, as well as to Lygaeus kalmii nymphal extract, and the response of L. kalmii nymphs towards O. fasciatus nymph extract was tested. It was found that there is an aggregation pheromone in the nymphs and young adults and that there may be an alarm pheromone in the old adult O. fasciatus. A preliminary chemical analysis performed on the aggregation pheromone revealed it probably to be a polar non-aromatic compound of high molecular weight.     

Flight fuel related differences between solitary and gregarious locusts (Locusta migratoria migratorioides)

Abstract. Flight fuel relations of crowded and isolated Locusta migratoria migratorioides were investigated in younger (12–16 days after fledging) and older (27–30 or 27–32 days after fledging) adult males.No phase polymorphism dependent differences were found in resting haemolymph carbohydrate levels of the younger locusts.In the older age group, resting haemolymph carbohydrate levels were slightly though significantly higher in the isolated than in the crowded locusts.Injection of various doses of synthetic adipokinetic hormones (AKHs) did not induce marked changes in haemolymph carbohydrate levels and no differences were found between crowded and isolated locusts.A 30 min flight led to the same decrease in haemolymph carbohydrate levels of isolated and crowded locusts, 43.3% and 44.6% of the resting levels, respectively.We concluded, therefore, that the results do not seem to indicate that isolated locusts rely more heavily on carbohydrates as flight fuel than crowded locusts.Hyperlipaemic responses to flight were less intense in isolated than in crowded locusts, but phase polymorphism dependent differences in flight-induced increase of haemolymph lipid levels were not parallel in 12–16-day-old and 27–32-day-old males.In the younger age group the difference was mainly in the duration of flight needed to induce full response which appeared already after 20 min of flight in the crowded locusts, but only after 45 or 60 min of flight in the isolated ones.In contrast, the older isolated locusts showed markedly lower haemolymph lipid elevations than the crowded locusts even after 30, 45 or 60 min of flight.The hypothesis is forwarded that isolated locusts have a rather coarse adipokinetic strategy focused on a single long-distance migratory flight, whereas gregarious locusts possess a fine adipokinetic balance for reiterative migratory flights and saving fuel reserves for unpredictable long-distance migrations.     

Albino corpus cardiacum extracts induce morphometric gregarization in isolated albino locusts, Locusta migratoria, that are deficient in corazonin

Abstract.  The neuropeptide [His⁷]-corazonin, present in the central nervous system and corpus cardiacum, is known to mimic a 'gregarizing' effect on phase-related morphometric ratios (hind femur length/maximum head width and fore wing length/hind femur length) when injected into locusts reared in isolation. However, an albino strain is known to exhibit phase-specific changes in these ratios in response to rearing density, although it is deficient in [His⁷]-corazonin. To examine whether there is a second factor responsible for this phenomenon, perhaps a corazonin-like factor that has lost its dark-colour inducing activity, methanol extracts of corpora cardiaca taken from crowd-reared albino nymphs of Locusta migratoria are injected into isolated-reared second-stadium albino nymphs and reared to adults in isolation. The hind femur length/maximum head width and fore wing length/hind femur length ratios are significantly different from those of control oil-injected counterparts, and shift significantly towards the values typical for crowd-reared gregarized individuals. The results indicate that the corpora cardiaca contain a factor similar to [His⁷]-corazonin, although it has no dark-colour inducing activity.     

Effects of isolation and crowding on the haemolymph pigment composition of the desert locust, Schistocerca gregaria (Orth., Acrididae)

Abstract:  An experimental study on the effect of isolation and crowding of the desert locust, Schistocerca gregaria (Forskål) on haemolymph pigment composition (as measured by absorbance ratios at 460 and 680 nm) was carried out at the International Centre of Insect Physiology and Ecology, Nairobi. The results showed that haemolymph absorbance ratios of gregarious and solitarious locusts were different (P < 0.05) and that there was no phase-specific sex differentiation with regard to this measure (P < 0.05). The haemolymph pigment composition changes during isolation and grouping showed contrasting trends. It changed rapidly in nymphs, but slowly and erratically in adults both resulting from isolation and grouping at hopper stage. Significant shifts in locust phase status were recorded in both situations in nymphs in F₀ generation, and in the adults by the end of the F₂ and F₃ generations of solitary and gregarious conditions, respectively. Shifting of fledglings did not induce significant changes in their mean absorbance ratios in both situations, i.e. isolation and crowding within F₀ generation. The ratio of length of posterior femur (F) to greatest width of the head capsule (C) changed either at a similar rate or more often slower than the haemolymph pigment ratio and both required more than a generation to transform fully. The study has shown that haemolymph pigment composition could be a suitable measure for monitoring phase changes at nymphal stages of the desert locust.     

Identification of

The neuropeptides inducing dark color in albino nymphs of the migratory locust Locusta migratoria were isolated from the larval brain of the silkworm, Bombyx mori and from the adult corpora cardiaca (CC) of the cricket Gryllus bimaculatus, respectively, and their amino acid sequences identified. The two peptides isolated from the two different species are identical to [Arg(7)] corazonin, a neuropeptide known to be present in a cockroach and others. This peptide induces a dark color in albino nymphs of L. migratoria at fmol levels, and a high dose of >/=100 pmol caused albino locusts to turn completely black, but it influenced neither body color nor metamorphosis in B. mori and G. bimaculatus. Therefore, the physiological functions of [Arg(7)] corazonin in the silkworm and the cricket remain unknown. The present study demonstrated the usefulness of the albino strain of L. mirgatoria as a specific bioassay system for this peptide.     

Locustol and its analogues

A new method of synthesis of the locust gregarization pheromone, locustol, is described, the pathway being from guaiacol to acetyl-isoacetovanillone, to isoacetovanillone, to 5-ethylguaiacol. These substances were all checked for their infrared spectra and were bioassayed. Only 5-ethylguaiacol affected chiasma frequency, hopper colour, and the $\text{F}\text{C}$ morphometric ratio, although guaiacol had some effect on chiasma frequency. Hopper and adult faeces, as well as grasshopper faeces, were steamextracted, tested by infrared spectrophotometry and by gas chromatography, and were bioassayed. Only the hopper faeces extract contained the pheromone as well as some isoacetovanillone. In the adult and grasshopper faeces extracts no aromatic substance could be found but the impurities contained, possibly, a mixture of a secondary alcohol, a carboxylic acid, and/or its ester, some of which also occurred in the impure hopper faeces extract. Isovanillin, 5-methylguaiacol, and amylacetate were also bioassayed: they showed no effect on chiasma frequencies, but the latter stimulated the production of a gregaria $\text{F}\text{C}$ ratio in solitaries. A figure is given of the chemical structures of eight substances which are part-analogues of locustol in regard to chiasma frequency, and of eight related substances which are chromosomally inactive but of which amylacetate stimulates the gregaria $\text{F}\text{C}$ ratio and methylformate stimulates the production of black hopper colour in solitaries. A consideration of these structures may aid in solving the chemical action of locustol.     

Knockdown of the corazonin gene reveals its critical role in the control of gregarious characteristics in the desert locust

The two plague locusts, Schistocerca gregaria and Locusta migratoria, exhibit density-dependent phase polyphenism. Nymphs occurring at low population densities (solitarious forms) are uniformly colored and match their body color to the background color of their habitat, whereas those occurring at high population densities (gregarious) develop black patterns. An injection of the neuropeptide, corazonin (Crz) has been shown to induce black patterns in locusts and affect the classical morphometric ratio, F/C (F, hind femur length; C, maximum head width). We herein identified and cloned the CRZ genes from S. gregaria (SgCRZ) and L. migratoria. A comparative analysis of prepro-Crz sequences among insects showed that the functional peptide was well conserved; its conservation was limited to the peptide region. Silencing of the identified SgCRZ gene in gregarious S. gregaria nymphs markedly lightened their body color and shifted the adult F/C ratio toward the value typical of solitarious forms. In addition, knockdown of the gene in solitarious nymphs strongly inhibited darkening even after a transfer to crowded conditions; however, these individuals developed black patterns after being injected with the Crz as a rescue treatment. SgCRZ was constitutively expressed in the brains of S. gregaria during nymphal development in both phases. This gene was highly expressed not only in the brain in both phases, but also in the corpora allata in the gregarious phase. This conspicuous phase-dependent difference in SgCRZ gene expression may indicate a functional role in the control of phase polyphenism in this locust.     

Mass spectrometric evidence for the deficiency in the dark-color-inducing hormone,.

A factor present in the brain and corpus cardiacum responsible for the induction of dark colour in Locusta migratoria was recently isolated and identified from the corpora cardiaca of normally pigmented locusts. The purification of this factor, designated as [His7]-corazonin was monitored using an albino mutant from a laboratory colony of an Okinawa (Japan) strain. In this study, we provide unequivocal mass spectrometric evidence that the brain and the corpora cardiaca of this albino Locusta mutant are deficient in [His7]-corazonin. Previously, [His7]-corazonin was shown to be responsible for the induction of dark body colour patterns as observed in crowded locusts. Using nanoflow-liquid chromatography-mass spectrometry, we demonstrated that this dark colour-inducing hormone is, however, present in the corpora cardiaca of solitary locusts (Schistocerca gregaria). Arch.