An analysis of the behavioural effects of crowding and re-isolation on solitary-reared adult desert locusts (Schistocerca gregaria) and their offspring

Abstract. The time-course of behavioural change in response to crowding and re-isolation was investigated in adults of Schistocerca gregaria Forskål (Orthoptera, Acrididae) using logistic regression analysis. Crowding solitary-reared adults for a period of 4 h caused them to behave similarly to crowd-reared insects, with their becoming much more active and moving towards rather than away from a stimulus group of locusts. Responsiveness to crowding was greatest in young adults. The behaviour acquired after 48 h of crowding was lost within 1 day of re-isolation. Although experience by solitary-reared adults of crowding for 48 h had only transitory effects on their own behaviour, there was also a long-term influence on the behaviour of their offspring. The strength of this effect was dependent on the age at which adults experienced crowding, increasing in a graded manner with adult age, and hence the recency of crowding before oviposition. Parents crowded at a late stage in the reproductive cycle yielded hatchlings which behaved indistinguishably from those from crowd-reared adults. Such an effect is consistent with the idea that females, through their previous experience of crowding, are effectively predicting the probability that their offspring will emerge into a high-density population, and predisposing their hatchlings' behaviour accordingly.     

Effects of crowding, isolation, and transfer from isolation to crowding on total ecdysteroid content of eggs in Schistocerca gregaria

We examined the ecdysteroid content of eggs from crowd-reared and solitary-reared desert locusts, Schistocerca gregaria, throughout embryogenesis from the day of egg laying until shortly before hatching on day 14. Depending on the time during incubation, ecdysteroid content in eggs from crowd-reared females was 5-10 times higher than in eggs from solitary-reared females. Our investigation revealed two peaks in ecdysteroid content of eggs from crowd-reared females, a small one at day 3 and a major peak at day 10 of incubation. At days 10, 12 and 14 during incubation of eggs from crowd-reared females, we found a positive correlation between egg mass and ecdysteroid content. There was no difference between eggs from the bottom and the top of individual egg pods, but variation in ecdysteroid content between egg pods from different females was considerable in all treatment groups. A brief crowding of solitary-reared females at the time of egg laying, a treatment that initiates maternally mediated gregarization of the developing offspring, had no effect on the consistently low ecdysteroid content in the eggs. This result rules out the possibility that the crowding experience of females is transmitted to the offspring by variation in the total amount of ecdysteroids in their eggs.     

Adult female desert locusts require contact chemicals and light for progeny gregarization

Crowding causes many organisms to express phenotypic plasticity in various traits. Phase polyphenism in desert locusts represents one extreme example in which a solitary form (solitarious phase) turns into a gregarious form (gregarious phase) in response to crowding. Conspicuous differences in body size and colour occur even in hatchlings. The phase‐specific differences in hatchling characteristics are caused by the tactile stimuli perceived by the antennae of their mother. However, the nature of the tactile stimuli and the mechanism by which the perceived stimuli are processed as a gregarizing signal remain unknown. To explore this problem, the antennae of solitarious adult females of the desert locust Schistocerca gregaria are touched with the bodies of conspecific locusts at different physiological stages and those of other species. The results suggest that a cuticular chemical factor at a specific developmental stage of conspecific locusts causes the solitarious females to produce large eggs that give rise to black hatchlings characteristic of gregarious forms (progeny gregarization), and that this or a similar compound occurs in other acridids, crickets and cockroaches but not in beetles. The involvement of a chemical substance is also supported by hexane extracts of cuticular surfaces of locusts that induce the same effects. Interestingly, crowding induces such gregarizing effects only when the female receives the appropriate stimulus in the presence of light. Solitarious female S. gregaria with their head capsule coated with phosphorescent paint exhibit progeny gregarization in response to crowding and light pulses in darkness, whereas those treated in the same way without light pulses fail to do so.     

Small-scale vegetation patterns in the parental environment influence the phase state of hatchlings of the desert locust

Desert locusts (Schistocerca gregaria Forskål (Orthoptera: Acrididae)) change phase in response to population density. Solitarious insects avoid one another; when crowded, they shift to the gregarious phase and aggregate. Laboratory experiments and individual‐based modelling have shown that small‐scale resource distribution can affect locust phase state via an influence on crowding. Laboratory work has also shown that parental phase state is transmitted to offspring via maternal inheritance. These effects had not been investigated in the field previously. We maintained small populations of adult desert locusts in semi‐field enclosures with different distribution patterns of a single plant species (Hyoscyamus muticus L. (Solanaceae)). The offspring of locusts exposed to more clumped patterns of vegetation exhibited more gregarious behaviour when tested in a behavioural phase assay than did progeny from parents left in enclosures with more scattered vegetation. These effects on nymphal behaviour appeared to be mediated by influences of resource distribution on adult phase state. Phase state in small semi‐field populations was influenced by small‐scale vegetation distribution. Phase differences engendered by environmental structure were maintained in time and transmitted to progeny.     

Paternal epigenetic effects of population density on locust phase‐related characteristics associated with heat‐shock protein expression

Many species exhibit transgenerational plasticity by which environmental cues experienced by either parent can be transmitted to their offspring, resulting in phenotypic variants in offspring to match ancestral environments. However, the manner by which paternal experiences affect offspring plasticity through epigenetic inheritance in animals generally remains unclear. In this study, we examined the transgenerational effects of population density on phase‐related traits in the migratory locust Locusta migratoria. Using an experimental design that explicitly controls genetic background, we found that the effects of crowd or isolation rearing on phase plasticity could be inherited to the offspring. The isolation of gregarious locusts resulted in reduced weight in offspring eggs and altered morphometric traits in hatchlings, whereas crowding of solitarious locusts exhibited opposite effects. The consequences of density changes were transmitted by both maternal and paternal inheritance, although the expression of paternal effects was not as pronounced as that of maternal effects. Prominent expression of heat‐shock proteins (Hsps), such as Hsp90, Hsp70 and Hsp20.6, could be triggered by density changes. Hsps were significantly upregulated upon crowding but downregulated upon isolation. The variation in parental Hsp expression was also transmitted to the offspring, in which the pattern of inheritance was consistent with that of phase characteristics. These results revealed a paternal effect on phase polyphenism and Hsp expression induced by population density, and defined a model system that could be used to study the paternal epigenetic inheritance of environmental changes.     

Transgenerational effects modulate density-dependent prophylactic resistance to viral infection in a lepidopteran pest

There is an increasing appreciation of the importance of transgenerational effects on offspring fitness, including in relation to immune function and disease resistance. Here, we assess the impact of parental rearing density on offspring resistance to viral challenge in an insect species expressing density-dependent prophylaxis (DDP); i.e. the adaptive increase in resistance or tolerance to pathogen infection in response to crowding. We quantified survival rates in larvae of the cotton leafworm (Spodoptera littoralis) from either gregarious- or solitary-reared parents following challenge with the baculovirus S. littoralis nucleopolyhedrovirus. Larvae from both the parental and offspring generations exhibited DDP, with gregarious-reared larvae having higher survival rates post-challenge than solitary-reared larvae. Within each of these categories, however, survival following infection was lower in those larvae from gregarious-reared parents than those from solitary-reared, consistent with a transgenerational cost of DDP immune upregulation. This observation demonstrates that crowding influences lepidopteran disease resistance over multiple generations, with potential implications for the dynamics of host–pathogen interactions.     

Density-dependent accumulation of phase characteristics in a natural population of the desert locust Schistocerca gregaria

Abstract. Phase characteristics of locusts from parents that experienced different population densities were investigated under field conditions in Morocco. The density experienced by adults induced a marked phase change in colour, behaviour and morphometry of their offspring. A high-density subpopulation gave rise to a preponderance of black hatchlings that exhibited a high level of aggregation as later stage nymphs and showed gregarious morphometric features as adults, whereas a low-density subpopulation produced a majority of green hatchlings with a lesser tendency to group as final-instar nymphs and more solitarious morphometry as adults. The constrained isolation of insects from the low-density subpopulation, or crowding of insects from the high-density subpopulation, resulted in a behavioural and morphometric change towards even more solitarious characteristics in the former and more pronounced gregarious characteristics in the latter, relative to field-caught insects of the same age. These results from the field are consistent with those in the laboratory and provide more evidence for the dual roles of an individual locust's experience of crowding as well as that of its parents in density-dependent phase change.     

Syntaxin 1A modulates the sexual maturity rate and progeny egg size related to phase changes in locusts

The migratory locust (Locusta migratoria) exhibits clear phenotypic plasticity depending on its population density. Previous studies have explored the molecular mechanisms of body colour, behavior, immunity, and metabolism between high population density gregarious (G) and low population density solitarious (S) locusts. However, the molecular mechanisms underlying differences in reproductive traits remain unknown. G locusts reach sexual maturation much faster and lay larger eggs compared with S locusts. The traits of G locusts decreased significantly with isolation, whereas those of S locusts increased with crowding. Analysis of gene expression in female adults indicated that syntaxin 1A (Syx1A) was expressed significantly higher in G locusts than in S locusts. After silencing Syx1A expression in G locusts by RNA interference (RNAi), their sexual maturity rate and progeny egg size changed towards those of S locusts. Similarly, increment in the traits of S locusts with crowding was blocked by Syx1A interference. Changes in the traits were also confirmed by decrease in the level of vitellogenin, which is regulated by Syx1A. In conclusion, plasticity of the sexual maturity rate and progeny egg size of G and S locusts, which is beneficial for locusts to adapt to environmental changes, is regulated by Syx1A.     

Fever and phenotype: transgenerational effect of disease on desert locust phase state

Natural enemy attack can cause transgenerational shifts in phenotype such that offspring are less vulnerable to future attack. Desert locusts (Schistocerca gregaria) show density‐dependent variation in their resistance to pathogens, such that they are less vulnerable to pathogens when in the high‐density gregarious phase state (when they would probably be more exposed to pathogens) than when in the solitarious phase state. We therefore hypothesized that infected gregarious parents would maintain this phenotype in their offspring. We infected gregarious desert locust nymphs with the fungal pathogen Metarhizium anisopliae var. acridum, and allowed them to survive to reproduction by means of behavioural fever. The phase state of the locust offspring was assessed by their colouration and behavioural assays. Contrary to our hypothesis, we found an increase in solitarization in the infected population (14.6% solitarious offspring from infected parents, vs. <2% from uninfected counterparts at equivalent density). In a second experiment, we simulated behavioural fever temperatures and obtained a similar result (13.6% solitarious offspring vs. 4.4% from controls), implying that the phenomenon is probably a side‐effect of the hosts’ fever response. Identification of this novel environmental factor affecting locust phase state could have important implications for the biological control of these major pests.     

The role of food distribution and nutritional quality in behavioural phase change in the desert locust

The behaviour of herbivorous insects is influenced by their nutritional state. Nutrition-induced behavioural changes are often interpreted as adaptive mechanisms for controlling nutrient intake; however, their influence on other life history traits has received far less attention. We investigated the effect of food quality and distribution on the behaviour and phase state of desert locusts, Schistocerca gregaria Forsk?l (Orthoptera, Acrididae), which change from the 'solitarious' to the 'gregarious' phase in response to population density. Phase change involves many morphological, physiological and behavioural changes. Solitarious insects are cryptic whereas gregarious locusts aggregate. Individual phase change is stimulated by mechanical contact with other locusts. A clumped resource distribution promotes change to the gregarious phase by increasing crowding and contact between individuals. In this study, we found that the effect of food distribution on locust phase depended on the nutritional quality of the food. We used three synthetic food treatments: near optimal, dilute and a choice of two unbalanced but complementary foods. Clumped resource distribution led to increased gregarization in the dilute and the complementary diet treatments. This effect was particularly pronounced on the complementary foods, owing to the interaction of crowding and locomotion. Gregarization was most pronounced in the dilute diet treatment, owing to increased activity. These diet-induced effects are explained in terms of behavioural changes in locomotion, quiescence and feeding that are consistent with what is known from earlier work on locust feeding behaviour and behavioural phase change. Copyright 2000 The Association for the Study of Animal Behaviour.     

A miniaturized assay to quantify effects of chemicals or physical stimuli upon locust activity

Solitary and gregarious locusts differ in many traits, such as body color, morphometrics and behavior. With respect to behavior, solitary animals shun each other, while gregarious animals seek each other＇s company, hence their crowding behavior. General activity, depending on the temperature, occurs throughout the day but is much lower in solitary locusts. Solitary locusts occasionally fly by night while gregarious locusts fly regularly during daytime （swarming）. In search of new assays to identify substances that control or modify aspects of （phase） behavior, we designed a simple activity assay, meant to complement existing behavioral measurement tools. The general activity is reflected in the number of wall hits, that is, the number of contacts between the locust and the vertical walls of a small arena. Using this single parameter we were able to quantify differences in total activity of both nymphs and adults of isolation-reared （solitary）, regrouped- and crowdreared （gregarious） locusts under different conditions. Furthermore, we demonstrate that there are inter- and intra-phase dependent differences in activities of 5th instar nymphs afar injections of the three different adipokinetic hormones.     

Immune responses of locusts to challenge with the pathogenic fungus Metarhizium or high doses of laminarin

Two isolates of Metarhizium anisopliae var acridum were tested for their effects on the locust immune system and for comparison with the effects of challenge by injection with laminarin. Isolate IMI 330189 (referred to hereafter as Met 189) is highly pathogenic whether applied topically as conidia or injected as blastospores. However, isolate ARSEF 728 (referred to hereafter as Met 728) is pathogenic only when injected as blastospores, suggesting that the lack of pathogenicity of topically applied conidia from this isolate is due to a failure to penetrate the insect cuticle and gain access to the haemocoel. After topical application of conidia from Met 189, no activation of prophenoloxidase is detected, but injection of blastospores from Met 189 brings about a transient increase in phenoloxidase activity in the haemolymph in both adult locusts and 5th instar nymphs, although this does not prevent fungal-induced mortality. Co-injection of adipokinetic hormone-I (AKH-I) with blastospores prolongs the activation of prophenoloxidase in the haemolymph of adult locusts, and enhances it in nymphs. It is argued that the lack of activation of prophenoloxidase in nymphs shown previously (Mullen, L., Goldsworthy, G., 2003. Changes in lipophorins are related to the activation of phenoloxidase in the haemolymph of Locusta migratoria in response to injection of immunogens. Insect Biochemistry and Molecular Biology 33, 661-670), reflects differences in the sensitivity of the immune system between adults and nymphs rather than distinct qualitative differences, and this is confirmed in this study by the demonstration that doses of laminarin higher than those used previously (>or=100 microg) do activate the prophenoloxidase cascade in 5th instar nymphs. Nodules are formed in locusts of all ages in response to fungal infection or injection of laminarin, although there is wide variation in the number, size and distribution of nodules formed. During the examination of 5th instar nymphs for nodule formation, a previously unknown phenomenon was observed in which the salivary glands melanise in response to injections of blastospores or high doses of laminarin. In c. 85% of such nymphs, this reaction is so strong that the whole salivary gland is intensely black. Such a response is not observed in the salivary glands of mature adult locusts.     

Genetic variation for parental effects on the propensity to gregarise in <Emphasis Type="Italic">Locusta migratoria</Emphasis>

Background  

Environmental parental effects can have important ecological and evolutionary consequences, yet little is known about genetic variation among populations in the plastic responses of offspring phenotypes to parental environmental conditions. This type of variation may lead to rapid phenotypic divergence among populations and facilitate speciation. With respect to density-dependent phenotypic plasticity, locust species (Orthoptera: family Acrididae), exhibit spectacular developmental and behavioural shifts in response to population density, called phase change. Given the significance of phase change in locust outbreaks and control, its triggering processes have been widely investigated. Whereas crowding within the lifetime of both offspring and parents has emerged as a primary causal factor of phase change, less is known about intraspecific genetic variation in the expression of phase change, and in particular in response to the parental environment. We conducted a laboratory experiment that explicitly controlled for the environmental effects of parental rearing density. This design enabled us to compare the parental effects on offspring expression of phase-related traits between two naturally-occurring, genetically distinct populations of Locusta migratoria that differed in their historical patterns of high population density outbreak events.     

Behavioural phase change in the Australian plague locust, Chortoicetes terminifera, is triggered by tactile stimulation of the antennae

Density-dependent phase polyphenism is a defining characteristic of the paraphyletic group of acridid grasshoppers known as locusts. The cues and mechanisms associated with crowding that induce behavioural gregarization are best understood in the desert locust, Schistocerca gregaria, and involve a combination of sensory inputs from the head (visual and olfactory) and mechanostimulation of the hind legs, acting via a transient increase in serotonin in the thoracic ganglia. Since behavioural gregarization has apparently arisen independently multiple times within the Acrididae, the important question arises as to whether the same mechanisms have been recruited each time. Here we explored the roles of visual, olfactory and tactile stimulation in the induction of behavioural gregarization in the Australian plague locust, Chortoicetes terminifera. We show that the primary gregarizing input is tactile stimulation of the antennae, with no evidence for an effect of visual and olfactory stimulation or tactile stimulation of the hind legs. Our results show that convergent behavioural responses to crowding have evolved employing different sites of sensory input in the Australian plague locust and the desert locust.     

Mermis nigrescens: physiological relationship with its host, the adult desert locust Schistocerca gregaria

Adult desert locusts were experimentally infected per os with 30, 50, or 60 Mermis nigrescens eggs, and changes in the host physiology were recorded. Larval nematodes were recovered from the hemocoel and counted at appropriate times after infection. The food consumption and blood volume of the host were unaffected by the parasitism, but the nematode significantly impaired the ability of male locusts to excrete the injected dye, amaranth, from their hemolymph. The total carbohydrate in the hemolymph of infected male and female locusts was severely depleted during the active growth period of the nematode and the possible utilization of these carbohydrates by the nematode are discussed. The total amino acid and protein levels in the blood of the host were unaffected by the nematode development, although concomittant changes in the levels of both these blood metabolites occurred in all locusts throughout the experimental period. However, although changes of this nature reflected the normal pattern of protein synthesis during oocyte development and oviposition in control locusts, the nematode suppressed oocyte development and caused oocyte resorption in the female host. The nematode did not significantly affect the level of total protein and amino acids in the flight muscles of male and female locusts, but a significant decrease in the level of fat body proteins and amino acids was recorded in infected hosts 16 and 21 days after infection. The possible effect of the nematode on protein metabolism by the host fat body is discussed in relation to the nutritional requirements of the nematode and involvement of the host endocrine system.     

The influence of environmental microstructure on the behavioural phase state and distribution of the desert locust Schistocerca gregaria

Abstract. In previous studies we used logistic regression analysis to quantify the change in behavioural phase state of Schistocerca gregaria (Forskål) nymphs subjected to variations in population density. Such work involved restricting insects in small containers either alone or in a crowd. In the present paper we have shown that the fine-scale distribution of food plants, perches and favourable microclimatic sites influences the spatial distribution of locusts, both in the laboratory and under semi-field conditions. When multiple resource sites were provided, solitarious locusts tended to disperse and behavioural gregarization was inhibited. However, provision of only a single site promoted congregation, overcoming the tendency of solitarious insects to avoid each other, and led to behavioural gregarization. The time-course and extent of this response was fully consistent with our earlier experiments using enforced crowding. We suggest that such quantitative, experimental studies of the effects of environmental microstructure on behaviour may yield fundamental insights into the dynamics of plague formation in the desert locust.     

Do desert locust hoppers develop gregarious characteristics by watching a video?

Various sensory stimuli have been suggested to induce gregarious body coloration in locusts, but most previous studies ignored the importance of substrate color. This study tested the effects of visual, olfactory and tactile stimuli from other locusts on the induction of gregarious body coloration in single (isolated-reared) Schistocerca gregaria nymphs housed in yellow-green cups. Odor from gregarious (crowd-reared) locusts, which is believed to induce black patterns in single locusts, had little effect when applied to visually isolated nymphs at the 2nd stadium onward, and all test nymphs remained green without black patterns at the last stadium, as in controls reared without odor and visual stimuli. Visual stimuli alone induced black patterns when a single solitarious nymph was allowed to see other locusts in another cup. The degree of black patterns increased as the number of locusts shown increased, and some test nymphs developed body coloration typically observed in gregarious forms. A classical morphometric ratio (hind femur length/head width) shifted toward the value typical of gregarious forms when the single nymphs were allowed to see 5 or 10 locusts. Single nymphs also developed black patterns when presented green conspecific nymphs and adults of two hemipteran species kept in another cup. No synergetic effects of visual and odor stimuli were detected. Movies of locusts, crickets and tadpoles were found effective in inducing black patterns in single locusts. Ontogenetic variation in the sensitivity to crowding and experimental methodology might be responsible for some discrepancies in the conclusions among different researchers.     

A behavioural analysis of phase change in the desert locust

A programme of research into phase change in the desert locust, Schistocerca gregaria, is described. The ability to change phase between solitarious and gregarious forms in response to population density is a key feature of locusts and is central to their occasional yet catastrophic impact on humans. Phase polymorphism is an extreme form of phenotypic plasticity. The most labile phase characteristic is behaviour. It is argued that a fully integrated study of behavioural phase change provides a powerful tool for understanding both the mechanisms of phase change and locust population dynamics, both of which offer possibilities for improved management and control of desert locust plagues. An assay for measuring behavioural phase-state in individual locusts was derived, based on logistic regression analysis. Experiments are described that used the assay to quantify the time-course of behavioural change, both within the life of individual locusts and across generations. The locust-related stimuli that provoke behavioural gregarization were investigated. Complex interactions were found between tactile, visual and olfactory stimuli, with the former exerting the strongest effect. Behavioural analysis also directed a study of the mechanisms whereby adult females exert an epigenetic influence over the phase-state of their developing offspring. Female locusts use their experience of the extent and recency of being crowded to predict the probability that their offspring will emerge into a high-density population, and alter the development of their embryos accordingly through a gregarizing agent added to the foam that surrounds the eggs at laying. There is also a less pronounced paternal influence on hatchling phase-state. An understanding of the time-course of behavioural phase change led to a study of the effect of the fine-scale distribution of resources in the environment on interactions between individual locusts, and hence on phase change. This, in turn, stimulated an exploration of the implications of individual behavioural phase change for population dynamics. Cellular automata models were derived that explore the relationships between population density, density of food resources and the distribution of resources in the environment. The results of the simulation showed how the extent of gregarization within a population increases with rising population size relative to food abundance and increasing concentration of food resources. Of particular interest was the emergence of critical zones across particular combinations of resource abundance, resource distribution and population size, where a solitarious population would rapidly gregarize. The model provided the basis for further laboratory and field experiments, which are described.     

The inhibitory effect of the fungal toxin, destruxin A, on behavioural fever in the desert locust

During an infection locusts behaviourally fever by seeking out higher environmental temperatures. This behaviour places the pathogen at sub-optimal growth temperatures while improving the efficiency of the immune system, thereby prolonging the lifespan of the host. It is therefore in the interest of the pathogen to either adapt to fever-like temperatures or to evolve mechanisms to interfere with, or inhibit fever. We investigated the behavioural fever response of desert locusts to two fungal pathogens. A prolonged fever was observed in locusts infected with Metarhizium acridum. However, fever was comparatively short-lived during infection with Metarhizium robertsii. In both cases restriction of thermoregulation reduced lifespan. Destruxin A (dtx A) produced by M. robertsii, but not M. acridum has previously been associated with the inhibition of the insect immune system. Injection of dtx A during infection with the fever-causing M. acridum inhibited fever and was particularly effective when administered early on in infection. Furthermore, locusts injected with dtx A were more susceptible to M. acridum infection. Therefore engineering M. acridum isolates currently used for locust biocontrol, to express dtx A may improve efficiency of control by interfering with fever.     

Body colour and genetic variation in winged morph production in the pea aphid

Aphids (Homoptera: Aphidoidea) produce a number of different phenotypes in their life-cycle, among which are winged (alate) and wingless (apterous) morphs. Lowe & Taylor (1964) and Sutherland (1969a, b) were the first to suggest that aphid clones differ in their propensity to produce the winged morph and that in the pea aphid (Acyrthosiphon pisum Harris), this propensity is linked to the colour of the phenotype. We tested for the occurrence of genetic variation in winged morph production by rearing individuals from red and green clones of pea aphid under wing-inducing (crowding) and control conditions, and scored the phenotypes of their offspring. Clones differed significantly in alate production and red clones produced on average a higher proportion of winged morphs than green clones. Importantly, however, there was considerable variation between clones of the same colour. Broad-sense heritabilities of winged morph production were 0.69 (crowding treatment) and 0.63 (control). Clones also differed in the number of offspring they produced. When exposed to the crowding stimulus, aphids deferred offspring production, resulting in a higher number of offspring produced in the crowding treatment than in the control.