Dianemobius Nigrofasciatus Research Articles

Interspecific competition leads to more long‐winged morphs in two sympatric cricket species

Abstract Coexistence mechanisms for species competing for the same resource include resource partitioning, neutrality, microhabitat preference and trade‐off between competitive and dispersal abilities. We explored the coexistence mechanism of two species of trigonidiid crickets (Dianemobius nigrofasciatus (Orthoptera: Trigonidiidae) and Polionemobius taprobanensis (Orthoptera: Trigonidiidae)) that share the same habitat. Dianemobius nigrofasciatus is more common in areas where the ground surface is somewhat open, while P. taprobanensis is more common in more densely vegetated environments. The effects of micro‐environmental differences, similarities in competitive ability and investment in dispersal ability under interspecific competitive conditions on the coexistence of these species were examined using laboratory experiments. Both P. taprobanensis and D. nigrofasciatus performed better in vegetated environments. Although the adult emergence of D. nigrofasciatus were delayed by the presence of P. taprobanensis, the emergence rate of P. taprobanensis was not significantly affected by D. nigrofasciatus. The presence of P. taprobanensis caused a higher frequency of a long‐winged morph (macropterous) of D. nigrofasciatus. The results suggest that D. nigrofasciatus is inferior to P. taprobanensis in interspecific competition, and it, therefore, disperses (as macropterous adults) at greater rates in the presence of P. taprobanensis. Furthermore, it may be that D. nigrofasciatus has been forced to change its preferred microhabitat from vegetative habitats, which are inherently more suitable, to more open environments due to competition. These above mechanisms are thought to allow the two species to coexist in the same habitat.

No evidence for the involvement of juvenile hormone III and 20-hydroxyecdysone in maternal decisions for embryonic diapause and diapause entry in the band-legged ground cricket Dianemobius nigrofasciatus (Orthoptera: Trigonidiidae)

No evidence for the involvement of juvenile hormone III and 20-hydroxyecdysone in maternal decisions for embryonic diapause and diapause entry in the band-legged ground cricket Dianemobius nigrofasciatus (Orthoptera: Trigonidiidae)

Differences in male calling song and female mate location behaviour between urban and rural crickets

Abstract Traffic noise is one of the selection pressures of urban habitats. Because noise interferes with acoustic communication, many studies have shown that the temporal structures of acoustic signals change depending on the noise. However, it is often unknown whether the changes are adaptive, such as changes being more easily located by females under noisy conditions. To investigate the phenomena, bend-legged ground crickets (Dianemobius nigrofasciatus) were collected from three urban and three rural habitats and reared in a common garden conditions. Male calling songs and female mate location behaviour of the progeny were measured. Urban males emitted a calling song with a higher dominant frequency and shorter chirp than did rural males. The playback experiment of female response and localization to songs of urban and rural males, controlling for noise, showed that there were no significant effects of the song on the mate location behaviour between urban and rural songs under noisy conditions. Although the song of urban males elicited a faster response of females, the effect was limited. These results indicate the change in song is not adaptive in terms of female choice. Urban females located more quickly to song under noisy conditions. The results suggest adaptive changes in female mating behaviour occur in urban habitats.

High incidence of leg autotomy in urban crickets

Abstract Predation is an important selection pressure affecting animals. Predation has both consumptive and non-lethal effects. Because it is often difficult to estimate the extent to which non-lethal effects occur, we focused on autotomy as an indicator of non-lethal effects occurring in urban habitats. Autotomy is expected to indicate that individuals have escaped from predators. The incidence of leg autotomy in the cricket Dianemobius nigrofasciatus was compared between urban and rural habitats. The incidence of autotomy was higher in urban than in rural habitats. Failure to moult and conspecific aggressive interactions were not the main factors contributing to the incidence of autotomy in these crickets. The ease of autotomy and risk-taking tendencies did not differ among habitats. These results indicate that the incidence of autotomy reflects predation failure, and the non-lethal effects of predation are more likely to occur in urban habitats.

The combined effects of artificial light at night and anthropogenic noise on life history traits in ground crickets

AbstractUrbanization affects various biotic and abiotic conditions through the disturbance and modification of the environment by human activity. Anthropogenic noise and artificial light at night (ALAN) associated with urbanization have negative impacts on wildlife. However, few studies have focused on the combined effects of noise and ALAN, and the counter‐adaptation of urban individuals to environmental pollution. We examined the single and combined effects of ALAN and noise on life history traits, including developmental period, survival rate, egg diapause, the number of progeny, and body size, of the cricket Dianemobius nigrofasciatus. We examined whether urban individuals are adapted to these factors by a common garden experiment using the crickets of both urban and rural origin. Our findings revealed no combined effect of ALAN and noise on the life‐history traits of D. nigrofasciatus except on that of survival and progeny production. Both ALAN and noise alone delayed development and prevented egg diapause. Neither noise nor ALAN had a significant effect on body size. The survival rate of rural crickets was not affected by noise and ALAN, but there was a negative synergetic effect. However, there were no significant effects of ALAN, noise, or their combined effect on the survival rate of urban crickets. These results suggest that urban individuals exhibit adaptive evolution to these factors. The number of progeny was positively affected by noise and not affected by ALAN, while the combined effect negatively affected the progeny production. The results suggest that a combined effect of noise and ALAN on important factors for the cricket such as survival and reproduction.

Phylomitogenomics of two Neotropical species of long-legged crickets Endecous Saussure, 1878 (Orthoptera: Phalangopsidae).

Mitochondrial genomes have provided significant insights into the evolution of several insects. A typical mitogenome contains 37 genes, and variations in gene order can indicate evolutionary relationships between species. In this study, we have assembled the first complete mitogenomes of Endecous chape and E. onthophagus and analyzed the phylogenetic implications for the Gryllidea infraorder. We performed DNA extractions and genome sequencing for both Endecous species. Subsequently, we searched for raw data in the Sequence Read Archive (SRA) in NCBI. Using the SRA data, we assembled the partial mitogenome of Dianemobius nigrofasciatus and annotated the protein-coding genes (PCGs) for nine species. Phylogenomic relationships were reconstructed using Maximum Likelihood (ML) and Bayesian Inference (BI), utilizing the PCGs from 49 Gryllidea species. The mitogenome lengths of E. chape and E. onthophagus are 16,266 bp and 16,023 bp, respectively, while D. nigrofasciatus has a length of 15,359 bp. Our results indicate that species within the infraorder exhibit four types of gene order arrangements that align with their phylogenetic relationships. Both phylogenomic trees displayed strong support, and the ML corroborated with the literature. Gryllidea species have significantly contributed to various fields, and studying their mitogenomes can provide valuable insights into this infraorder evolution.

The circadian clock gene (Clock) regulates photoperiodic time measurement and its downstream process determining maternal induction of embryonic diapause in a cricket

In response to short days in autumn, most temperate multivoltine insects enter diapause, a state in which development or reproduction is suppressed or arrested, which serves to coordinate their development and physiology (or that of offspring) with annual changes in the environment (i.e. photoperiodism). This response is mediated by a measurement of time based on photoperiod (photoperiodic time measurement), which is thought to be regulated by a circadian clock. However, some studies also demonstrate the involvement of the circadian clock in an output process that generates phenotypes associated with diapause. To gain further insight into this, we silenced the Clock (Clk) gene, the main regulator of the circadian clock, in the band-legged ground cricket Dianemobius nigrofasciatus (Orthoptera: Trigonidiidae). Silencing the Clk gene using RNA interference (Clk RNAi) in female crickets resulted in abnormal circadian rhythms under constant darkness and light-dark conditions, thereby indicating the central role of this gene in the circadian clock mechanism. Clk RNAi females exhibited long-day oviposition behaviour, even when reared under short-day conditions, thereby indicating the involvement of Clk in photoperiodic time measurement. In addition, Clk RNAi females immediately laid non-diapause-type eggs, which was not recorded in control females under either short-day or long-day conditions and cannot be explained in terms of dysfunction of photoperiodic time measurement. Accordingly, we speculate that Clk could also be involved in a downstream process that results in the laying of diapause-type eggs.

Effect of diet on body size and survival of omnivorous crickets

AbstractOmnivory is a feeding habit in which animals prey on organisms from two or more trophic levels. Omnivorous species play important roles in predator–prey interactions, food web dynamics, and ecosystem functions. Although cricket species have been considered omnivores, quantified investigations of their feeding habits have not been carried out. Therefore, we compared the developmental time, body size and mortality rate of the band‐legged ground cricket, Dianemobius nigrofasciatus (Matsumura) (Orthoptera: Trigonidiidae) reared on diets meant for carnivores and herbivores. Furthermore, to examine whether the crickets can balance their nutritional conditions through their food choices, we set up two additional diet treatments, which comprised diets for carnivores and herbivores provided simultaneously or by rotation. The mortality rate of the carnivore diet treatment was higher than that of the other diet treatments. This result suggests that D. nigrofasciatus is a “herbivore” that also obtains nutrients from resources other than plant materials. Although developmental time did not significantly differ among the dietary treatments, the body size of the crickets that were provided both diets simultaneously was significantly larger than that of the crickets provided with only the diets for either carnivores or herbivores. These results suggest that an appropriate amount of carnivorous feeding is advantageous for the development of the cricket, and indicates that crickets can balance nutrition on their own.

Robustness of latitudinal life-cycle variations in a cricket Dianemobius nigrofasciatus (Orthoptera: Trigonidiidae) in Japan against climate warming over the last five decades

Recent climate warming is expected to have changed the phenology of organisms. The cricket Dianemobius nigrofasciatus (Matsumura) (Orthoptera: Trigonidiidae) has univoltine and bivoltine life cycles in northern and southern regions of Japan, respectively, because of latitudinal variation in the growing season length. Its adult body size increases with decreasing latitude, and decreases at a latitude where the number of annual generations increases. The present study aims to examine whether the range of the bivoltine life cycle has expanded northward due to climate warming. We compared a latitudinal saw-tooth body size cline between the 1960s and the 2010s. The body size showed a latitudinal saw-tooth cline in adults collected in recent years, as it did in adults collected five decades ago. However, no significant difference was observed between these two clines, suggesting that the bivoltine life cycle has not expanded in the last five decades. These results contrast to those recently reported in a closely related species, Polionemobius mikado (Shiraki) (Orthoptera: Trigonidiidae), in which the turning points of the saw-tooth cline had shifted northward in the last four decades. The stable latitudinal distribution of life cycles in D. nigrofasciatus might result from the different photoperiodic regulation of growth rate involved in the different responses to climate warming.

Cell cycle regulator, small silencing RNA, and segmentation patterning gene expression in relation to embryonic diapause in the band-legged ground cricket

Insects enter diapause to synchronize their life cycle with biotic and abiotic conditions favorable for their development, reproduction, and survival. Adult females of the band-legged ground cricket Dianemobius nigrofasciatus (Orthoptera, Glyllidae) respond to environmental factors in autumn and lay diapause-destined eggs. The eggs arrest their development and enter diapause at a very early embryonic stage, specifically the cellular blastoderm. To elucidate the physiological mechanisms underlying this very early stage programmed developmental arrest, we investigated the cell division cycle as well as the expression of cell cycle regulators, small silencing RNAs, and segment patterning genes. The diapause embryo arrests its cell cycle predominantly at the G0/G1 phase. The proportion of cells in the S phase of the cell cycle abruptly decreased at the time of developmental arrest, but further changes of the G0/G1 and G2/M were later observed. Thus, cell cycle arrest in the diapause embryo is not an immediate event, but it takes longer to reach the steady state. We further elucidated molecular events possibly involved in diapause preparation and entry. Downregulation of Proliferating cellular antigen (PCNA; a cell cycle regulator), caudal and pumilio (cad and pum; early segmentation genes) as well as P-element induced wimpy testis (piwi) (a small silencing RNA) prior to the onset of developmental arrest was notable. The downregulation of PCNA, cad and pum continued even after entry into developmental arrest. In contrast to upregulation in non-diapause eggs, Cyclin D (another cell cycle regulator) and hunchback, Krüppel, and runt (gap and pair-rule genes) were downregulated in diapause eggs. These molecular events may contribute to embryonic diapause of D. nigrofasciatus.

Abiotic effects on the distributions of major insect species in agricultural fields

We compared how abiotic factors affect the regional distributions of four insect groups (Dermaptera, Formicidae, Orthoptera and Diptera). Insects were collected using a pitfall trap from 36 agricultural field sites, and 31 933 individuals encompassing 139 species were observed. The distribution pattern of dominant species was not obviously different among the four groups. Species richness (H′) of Orthoptera showed a negative and positive correlation with precipitation and sunlight duration, respectively. Longitudinal zonation was the main distribution pattern of many dominant species. Overall, canonical correspondence analysis showed that temperature and precipitation were closely associated with the distribution of Orthoptera, but not that of Formicidae, implying that Formicidae may be affected less by abiotic factors compared to other taxonomic groups. Some orthopteran species such as Dianemobius nigrofasciatus and Polionemobius mikado showed a negative correlation with temperature. We suggest that these species can be categorized as susceptible to climate change. Our results also implied that the association between climate variables and distribution of insects should be evaluated at a species level.

Ground crickets singing in volcanic warm “islets” in snowy winter: Their seasonal life cycles, photoperiodic responses and origin

AbstractThe ground cricket Dianemobius nigrofasciatus overwinters as an egg in Japan, being univoltine in Hokkaido and northern Honshu and bivoltine farther south. In Hokkaido, however, this cricket is heard singing in winter in several fumarolic fields covered with moss and grasses locally known as “bokke”. In such warm “islets” the adult density was high in early summer and again in autumn, indicating that the cricket is bivoltine in contrast to the univoltine life cycle outside the bokke habitats in Hokkaido. Eggs laid by females collected at regular intervals from a bokke habitat showed a clear seasonal cycle of diapause incidence. At 26°C, the bokke strains produced non‐diapause eggs under long days and diapause eggs under short days as in the southern bivoltine populations, although the critical day‐length was longer than in the south. Several strains derived from non‐bokke habitats in Hokkaido and northern Honshu produced high percentages of diapause eggs under long days as well as short days as expected for the univoltine life cycle. Winter adults singing in bokke habitats could be either survivors of the autumn generation or individuals derived from eggs laid in autumn and then matured in response to the high soil temperature. In the laboratory, the proportion of egg diapause in short days was decreased by selection only for several generations. Phylogenetic trees of bokke and non‐bokke populations, based on both the nucleotide sequence of the mitochondrial COI gene and four allozyme loci, suggest that bokke populations have not been isolated from non‐bokke populations for an evolutionarily significant time.

Effect of hind-limb autotomy on calling and hiding behavior in the band-legged ground cricket, Dianemobius nigrofasciatus

Crickets can escape death by autotomizing a limb when attacked by predators. In contrast with this benefit, autotomized individuals pay an immediate cost of escape speed and mating ability. Therefore, an adaptive response compensating for the cost of autotomy might be advantageous in autotomized individuals. In this study, we examined whether autotomy induced behavioral plasticity compensating for future cost in the band-legged ground cricket Dianemobius nigrofasciatus. Behavioral traits of D. nigrofasciatus were compared between autotomized and intact individuals. Frequency of calling behavior was higher for autotomized males. This behavior might be advantageous because females prefer actively calling males. In contrast with calling behavior, the frequencies of hiding behavior did not vary between autotomized and intact crickets, irrespective of sex. It might be disadvantageous for both sexes to hide, because females could not find hiding males and hiding females could not find males. These results indicated autotomy-induced behavioral plasticity that might reduce the cost of autotomy.

Serotonin- and two putative serotonin receptors-like immunohistochemical reactivities in the ground crickets Dianemobius nigrofasciatus and Allonemobius allardi

Serotonin (5-hydroxytryptamine; 5-HT)- and two putative serotonin receptors, 5-HT1A- and 5-HT1B-like, immunohistochemical reactivities were investigated in the cephalic ganglia of two ground crickets, Dianemobius nigrofasciatus and Allonemobius allardi. 5-HT-ir was strongly expressed in the central body, accessory medulla region of the optic lobe, frontal ganglion, posterior cortex of the protocerebrum, dorsolateral region of the protocerebrum, and the suboesphageal ganglion (SOG) in both crickets. However, 5-HT1A-ir and 5-HT1B-ir showed quite mutually distinct patterns that were also distinct from 5-HT-ir. 5-HT1A-ir was located in the pars intercerebralis, dorsolateral region of the protocerebrum, optic tract, optic lobe, and the midline of the SOG in both crickets. 5-HT1B-ir was located in the pars intercerebralis and dorsolateral region of the protocerebrum, and detected weakly in the optic lobe, tritocerebrum, and the midline of the SOG in both crickets. Interspecific differences were observed with 5-HT1A-ir. 5-HT1A-ir was expressed weakly in two neurons in the mandibular neuromere of the SOG in D. nigrofasciatus, while it was expressed strongly in the tritocerebrum, mandibular neuromere, and maxillary neuromere of the SOG in A. allardi and co-localized with CLOCK-ir (CLK-ir). 5HT-1B-ir was co-localized with CLK-ir in the tritocerebrum, mandibular neuromere, and maxillary neuromere of the SOG when double-labeling was conducted in both crickets. These results indicated that 5-HT and both types of 5-HT receptors may regulate circadian photo-entrainment or photoperiodism in A. allardi, while only 5-HT1B may be involved in circadian photo-entrainment or photoperiodism in D. nigrofasciatus.

Autotomy-induced life history plasticity in band-legged ground cricketDianemobius nigrofasciatus

Crickets can autotomize their limbs when attacked by predators. This enables them to escape death, but imposes a short-term cost on their escape speed and a long-term cost on their future mating ability. Therefore, adaptive response compensated for the cost of autotomy might be advantageous for autotomized individuals. In the present study, we examined whether autotomy induced life history plasticities compensating for the future cost in the band-legged ground cricket Dianemobius nigrofasciatus. Life history traits of D. nigrofasciatus were compared between autotomized and intact individuals. The developmental time and head width of the individuals that were autotomized as fourth instar nymphs were significantly shorter and smaller, respectively, than those of intact individuals. However, the adult longevity, number of eggs laid and oviposition schedule did not vary between autotomized and intact individuals. In addition, there was no difference between individuals autotomized at the fourth instar and adult stages in these three traits. Early maturation in the autotomized individuals might be advantageous through reducing the risk of predation owing to the shorter period in nymphal stages. The cost of small body size in the autotomized females might not be so great because of no significant difference in fecundity between autotomized and intact individuals. However, the cost of small body size was unclear in the autotomized males because in general larger males were preferred by females. These results indicated autotomy-induced life history that might reduce the cost of autotomy.

Early embryonic development and diapause stage in the band-legged ground cricket Dianemobius nigrofasciatus

The band-legged ground cricket Dianemobius nigrofasciatus enters diapause at an early embryonic stage when adults are reared under short-day conditions or the eggs are exposed to a low temperature. We examined the morphological features of the embryo during early development and determined the exact stage of entry into diapause. In non-diapause eggs, no periplasmic space was observed in the surface region and a small number of nuclei surrounded by cytoplasm (energids) were found among the yolk granules and lipid droplets 12 h after egg laying (AEL) at 25 degrees C. The energids sparsely but evenly populated the surface region at 40 h AEL, but there were some gaps between these energids. A continuous thin layer of nuclei with cytoplasm had completely covered the egg surface at 56 h AEL, suggesting that the blastoderm is formed between 40 and 56 h AEL. At 72 h AEL, we found a germ band at the posterior pole. Electron microscopy revealed clear cell membranes at 40 h AEL. Staining with rhodamine-dextran dye demonstrated that the cell membrane is formed when the nuclei appear on the egg surface at 12-24 h AEL. These results indicate that cellularization occurs before blastoderm formation. In diapause eggs, neither the embryonic rudiment nor germ band was formed, but a continuous layer of cells covered the egg surface. It is concluded that D. nigrofasciatus enters diapause at the cellular blastoderm.

Characterization of arylalkylamine N-acetyltransferase (AANAT) activities and action spectrum for suppression in the band-legged cricket, Dianemobius nigrofasciatus (Orthoptera: Gryllidae)

Arylalkylamine N-acetyltransferase (AANAT), constituting a large family of enzymes, catalyzes the transacetylation from acetyl-CoA to monoamine substrates, although homology among species is not very high. AANAT in vertebrates is photosensitive and mediates circadian regulation. Here, we analyzed AANAT of the cricket, Dianemobius nigrofasciatus. The central nervous system contained AANAT activity. The optimum pHs were 6.0 (a minor peak) and 10.5 (a major peak) with crude enzyme solution. We analyzed the kinetics at pH 10.5 using the sample containing collective AANAT activities, which we term AANAT. Lineweaver-Burk plot and secondary plot yielded a K(m) for tryptamine as substrate of 0.42 microM, and a V(max) of 9.39 nmol/mg protein/min. The apparent K(m) for acetyl-CoA was 59.9 microM and the V(max) was 8.14 nmol/mg protein/min. AANAT of D. nigrofasciatus was light-sensitive. The activity was higher at night-time than at day-time as in vertebrates. To investigate most effective wavelengths on AANAT activity, a series of monochromatic lights was applied (350, 400, 450, 500, 550, 600 and 650 nm). AANAT showed the highest sensitivity to around 450 nm and 550 nm. 450 nm light was more effective than 550 nm light. Therefore, the most effective light affecting AANAT activity is blue light, which corresponds to the absorption spectrum of blue wave (BW)-opsin.

Maternal control of cold and desiccation tolerance in eggs of the band‐legged ground cricket Dianemobius nigrofasciatus in relation to embryonic diapause

AbstractCold and desiccation tolerance was investigated in the eggs of the band‐legged ground cricket Dianemobius nigrofasciatus in relation to embryonic diapause. Diapause eggs were more tolerant to both desiccation and cold than non‐diapause eggs. In addition, diapause‐destined eggs on day zero (0–12 h after being laid) already showed high tolerance to these stresses before entering diapause. This clearly indicates that stress tolerance, like diapause, is controlled by photoperiod, but is not directly associated with diapause itself. Because the acquisition of stress tolerance predates the onset of diapause, it is plausible that diapause programming during some period before the onset of diapause is involved in the acquisition of stress tolerance. Weights and sizes were nearly identical in short‐day and long‐day eggs until day five. Sorbitol, a major sugar alcohol in eggs of D. nigrofasciatus, was accumulated at the same level in short‐day and long‐day eggs on days zero and five. These results indicate that the surface‐to‐volume ratio as well as the accumulation of sugar alcohol is not involved in the acquisition of stress tolerance. Maternal factors are clearly involved in the acquisition of stress tolerance in D. nigrofasciatus eggs, but the physiological mechanisms underlying the tolerance are still unclear.

Molecular structure, expression patterns, and localization of the circadian transcription modulator CYCLE in the cricket, Dianemobius nigrofasciatus

CYCLE (CYC), also known as BMAL1 in vertebrate nomenclature, is a transcription modulator of the circadian genes period and timeless of Drosophila melanogaster. We cloned a cDNA encoding a CYC homologue from the head of the ground cricket, Dianemobius nigrofasciatus (Dncyc), the first CYC from Hemimetabola. The deduced sequence corresponded to a 601 amino-acid polypeptide, with well-defined bHLH, PAS-A, PAS-B, PAC, and BTCR domains. The amino-acid sequence showed 70.7% identity with the CYC protein of Athalia rosae, 63.8% with D. melanogaster, and 52% identity with the human homologue. A cyc transcript of around 3.6kb occurs in the brain, midgut, testis, fatbody, and muscle. An additional band of around 1.1kb gave a hybridization signal in the head. No temporal oscillation in cyc mRNA abundance was observed in the head of the adult cricket when investigated by Northern blot analysis. CYC-like immunohistochemical reactivity (ir) and its dimerization partner CLOCK (CLK)-ir appeared in the pars intercerebralis (PI), tritocerebrum, dorsolateral protocerebrum, and subesophageal ganglion (SOG), but no CYC-ir was observed in the optic lobe (OL) that showed CLK-ir. The deutocerebrum showed a unique CLK-ir but no CYC-ir pattern. Double-labelling experiments showed that both antigens were co-localized in the mandibular and maxillary neuromeres of the SOG. CYC-ir showed no daily oscillation in intensity and the staining pattern was always cytoplasmic. CLK-ir occurred in the nucleus at ZT 16, but was cytoplasmic at other ZT times. A neuronal network equivalent to adult system occurred in the second nymphal stadium.

Neurohormones as putative circadian clock output signals in the central nervous system of two cricket species

Antisera to the neuropeptides corazonin (Crz) and crustacean cardioactive peptide (CCAP) and to the diapause hormone (DH) react with small sets of neurones in the cephalic ganglia of the crickets Dianemobius nigrofasciatus and Allonemobius allardi. The distribution of their immunoreactivities is similar in the two species and overlaps with the locations of presumed circadian clock components in the optic lobes, protocerebrum, tritocerebrum, suboesophageal ganglion (SOG) and frontal ganglion. D. nigrofasciatus contains two Crz-immunoreactive (Crz-ir) cells in each optic lobe, six cell groups in the protocerebrum, four in the tritocerebrum, and one in SOG, whereas A. allardi harbours only five Crz-ir groups in the protocerebrum and four in the tritocerebrum. CCAP immunoreactivity occurs in both species in four protocerebrum cell clusters, four tritocerebrum cell clusters, four SOG cell clusters, one frontal ganglion cell cluster, and two optic lobe cell clusters; D. nigrofasciatus possesses two additional cells with unique links to the lamina in the optic lobe. DH-related antigens are present in four cell clusters in the optic lobe, six (D. nigrofasciatus) or eight (A. allardi) in the protocerebrum, four in the tritocerebrum, and three (A. allardi) or five (D. nigrofasciatus) in the SOG. Some of the detected cells also react with antibody to the clock protein Period (PER) or lie close to PER-ir cells. Crickets reared at two different photoperiods do not differ in the distribution and intensity of immunoreactivities. No changes have been detected during the course of diurnal light/dark cycles, possibly because the antisera react with persistent prohormones, whereas circadian fluctuations may occur at the level of their processing or of hormone release. The projection of immunoreactive fibres to several brain regions, the stomatogastric nervous system and the neurohaemal organs indicates multiple functions of the respective hormones.