Riptortus Pedestris Research Articles

Binding properties of chemosensory protein 12 in Riptortus pedestris to aggregation pheromone (E)-2-hexenyl (Z)-3-hexenoate

Riptortus pedestris (bean bug), a common soybean pest, has a highly developed olfactory system to find hosts for feeding and oviposition. Chemosensory proteins (CSPs) have been identified in many insect species; however, their functions in R. pedestris remain unknown. In this study, quantitative real time-polymerase chain reaction (qRT-PCR) revealed that the expression of RpedCSP12 in the adult antennae of R. pedestris increased with age. Moreover, a significant difference in the expression levels of RpedCSP12 was observed between male and female antennae at one and three days of age. We also investigated the binding ability of RpedCSP12 to different ligands using a prokaryotic expression system and fluorescence competitive binding assays. We found that RpedCSP12 only bound to one aggregation pheromone, (E)-2-hexenyl (Z)-3-hexenoate, and its binding decreased with increasing pH. Furthermore, homology modelling, molecular docking, and site-directed mutagenesis revealed that the Y27A, L74A, and L85A mutants lost their binding ability to (E)-2-hexenyl (Z)-3-hexenoate. Our findings highlight the olfactory roles of RpedCSP12, providing insights into the mechanism by which RpedCSPs bind to aggregation pheromones. Therefore, our study can be used as a theoretical basis for the population control of R. pedestris in the future.

Leafhopper transmits soybean stay-green associated virus to leguminous plants

A novel geminivirus, soybean stay-green associated virus (SoSGV), was previously shown to cause soybean delayed senescence and is associated with the incidence of soybean stay-green syndrome. The modes of SoSGV transmission were not yet known. We captured insects belonging to 24 distinct species in a soybean field with the SoSGV outbreak and detected the presence of SoSGV only in leafhoppers and bean bugs (Riptortus pedestris). Caged feeding experiments using captured leafhoppers and bean bugs from soybean fields showed that leafhoppers, but not bean bugs, are vectors transmitting SoSGV. The common brown leafhopper (Orosius orientalis) is identified as the dominant leafhopper species and can establish colonies feeding on soybean plants in experimental conditions. An investigation of SoSGV defective DNA revealed that soybean genomic DNA fragments could be inserted into the SoSGV genome, while sequences from wild soybean, red bean, and cowpea were also identified. We further showed that the common brown leafhopper could transmit SoSGV to wild soybean and red bean plants, emphasizing a vector’s role of the leafhopper in the transmission of SoSGV in the field.

A secreted salivary effector from Riptortus pedestris impairs soybean defense through modulating phytohormone signaling pathways.

Riptortus pedestris (Fabricius), one of the major piercing-sucking insects in soybeans, causes delayed plant senescence and abnormal pods, known as staygreen syndrome. Recent research has shown that direct feeding of this insect is the major cause of soybean staygreen syndrome. However, it remains unclear whether R. pedestris salivary proteins play vital roles in insect infestation. Here, we found that 4 secretory salivary proteins can induce cell death in Nicotiana benthamiana by transient heterologous expression. The cell death induced by Rp2155 relies on the nucleotide-binding leucine-rich repeat helper, HSP90. Tissue-specificity assays indicated that Rp2155 is specifically expressed in the salivary gland of R. pedestris and is significantly induced during insect feeding. The expression of salicylic acid (SA)-, jasmonic acid (JA)-related genes was increased in soybean when fed by Rp2155-silenced R. pedestris. More importantly, soybean staygreen symptoms caused by R. pedestris were significantly alleviated when Rp2155 was silenced. Together, these results suggest that the salivary effector Rp2155 is involved in promoting insect infestation by suppressing the JA and SA pathways, and it can be considered as a potential RNA interference target for insect control.

The gut bacterium Serratia marcescens mediates detoxification of organophosphate pesticide in Riptortus pedestris by microbial degradation

AbstractOrganophosphate insecticides have been widely used in agriculture to control insect pests and raised growing problems including insecticide resistance. According to recent studies, microbial symbionts play important roles in developing insecticide resistance. The bean bug Riptortus pedestris is a notorious pest of leguminous crops, harbouring diverse microorganisms in their insect midguts. However, the potential biological roles of many gut‐associated bacteria are largely unknown. In this study, we isolated a bacterial strain from the midgut of R. pedestris and identified it as Serratia marcescens based on its morphological characteristics and the phylogenetic analysis of the 16S rRNA gene sequence. A halo of phosphorus‐solubilizing bacteria was observed on a medium containing lecithin and an organophosphorus insecticide dimethoate. Subsequently, the microbial degradation of dimethoate was determined using Gas Chromatography with Mass Spectroscopy (GC–MS). Further analysis indicated that the S. marcescens contained organophosphorus‐degrading MBL‐fold metallo‐hydrolase gene. Additionally, we discovered that the bacterium could colonize the insect midgut stably and enhance the host survivorship when exposed to dimethoate. Our results revealed that an insect gut‐associated bacterium could degrade organophosphorus compounds, thereby reducing related insecticide toxicity to the insect hosts. These findings provided further insights into the host‐microbiota interactions underpinning the development of insecticide resistance.

Identification and characterization of gut-associated lactic acid bacteria isolated from the bean bug, Riptortus pedestris (Hemiptera: Alydidae).

Lactic acid bacteria (LAB) are beneficial bacteria for humans and animals. However, the characteristics and functions of LAB in insects remain unclear. Here, we isolated LAB from the gut of Riptortus pedestris, a pest that is a significant problem in soybean cultivation in Korea, and identified two Lactococcus lactis and one Enterococcus faecalis using matrix-associated laser desorption/ionization-time of flight and 16S rRNA analyses. All three LAB strains survived at pH 8, and L. lactis B103 and E. faecalis B105 survived at pH 9 for 24 h. In addition, these strains survived well in simulated gastric juice of humans containing pepsin and exhibited high resistance to bile salts. Two strains of L. lactis and one of E. faecalis maintained constant density (> 104 colony-forming units [CFU]/mL) at pH 2.5, but viability at pH 2.2 was strain-dependent. The three LAB were reinoculated into second-instar nymphs of R. pedestris and colonized well, reaching a constant density (> 105 CFU/gut) in the adult insect gut. Interestingly, feeding of these LAB increased the survival rate of insects compared to the negative control, with the largest increase seen for L. lactis B103. However, the LAB did not increase the weight or length of adult insects. These results indicate that insect-derived LAB possess the traits required for survival under gastrointestinal conditions and have beneficial effects on insect hosts. The LAB infection frequency of the wild bean bug populations was 89% (n = 18) in Gyeongsangnam-do, South Korea. These LAB can be utilized as a novel probiotic in the cultivation of beneficial insects. This study provides fundamental information about the symbiosis between insects and LAB, and a novel concept for pest control.

Feeding Behavior Comparison of Bean Bugs, Riptortus pedestris and Halyomorpha halys on Different Soybean Cultivars

Riptortus pedestris (Fabricius) and Halyomorpha halys (Stål) are the major pests that feed on soybean pods, seeds, and fruits. Higher populations and damage occur during the soybean maturity stages (podding to harvest). To compare the feeding behavior of R. pedestris and H. halys, we used the six most cultivated cultivars (Daepung-2ho, Daechan, Pungsannamul, Daewon, Seonpung, and Seoritae) in Korea using the electropenetrography (EPG) technique. Both R. pedestris and H. halys, the NP (non-penetration), a non-probing waveform, was the shortest in the Pungsannamul (298 and 268 min) and the longest in the Daepung-2ho (334 and 339 min), respectively. The feeding waveforms Pb (phloem feeding: E1-Salivation and E2-Sap feeding) and G (xylem feeding) were the longest in Pungsannamul and the shortest in Daepung-2ho. In addition, as a result of investigating the damage rate by planting six cultivars of beans in the field, as expected, the proportions of damage types B and C were highest in Pungsannamul and lowest in Daepung-2ho. These results reveal that both bug species ingest xylem sap from leaflets and stems using a salivary sheath strategy to acquire water and nutrients from soybean pods/seeds with cell-rupture tactics. This study provides perceptive information to understand the feeding behavior, field occurrence, and damage patterns of R. pedestris and H. halys, which may have key implications for the management of hemipteran pests by determining the specificity and susceptibility of host plants.

A salivary secretory protein from Riptortus pedestris facilitates pest infestation and soybean staygreen syndrome.

The bean bug (Riptortus pedestris), one of the most important pests of soybean, causes staygreen syndrome, delaying plant maturation and affecting pod development, resulting in severe crop yield loss. However, little is known about the underlying mechanism of this pest. In this study, we found that a salivary secretory protein, Rp614, induced cell death in nonhost Nicotiana benthamiana leaves. NbSGT1 and NbNDR1 are involved in Rp614-induced cell death. Tissue specificity analysis showed that Rp614 is mainly present in salivary glands and is highly induced during pest feeding. RNA interference experiments showed that staygreen syndrome caused by R. pedestris was significantly attenuated when Rp614 was silenced. Together, our results indicate that Rp614 plays an essential role in R. pedestris infestation and provide a promising RNA interference target for pest control.

Detection of Soybean Insect Pest and a Forecasting Platform Using Deep Learning with Unmanned Ground Vehicles

Soybeans (Glycine max (L.) Merr.), a popular food resource worldwide, have various uses throughout the industry, from everyday foods and health functional foods to cosmetics. Soybeans are vulnerable to pests such as stink bugs, beetles, mites, and moths, which reduce yields. Riptortus pedestris (R. pedestris) has been reported to cause damage to pods and leaves throughout the soybean growing season. In this study, an experiment was conducted to detect R. pedestris according to three different environmental conditions (pod filling stage, maturity stage, artificial cage) by developing a surveillance platform based on an unmanned ground vehicle (UGV) GoPro CAM. Deep learning technology (MRCNN, YOLOv3, Detectron2)-based models used in this experiment can be quickly challenged (i.e., built with lightweight parameter) immediately through a web application. The image dataset was distributed by random selection for training, validation, and testing and then preprocessed by labeling the image for annotation. The deep learning model localized and classified the R. pedestris individuals through a bounding box and masking in the image data. The model achieved high performances, at 0.952, 0.716, and 0.873, respectively, represented through the calculated means of average precision (mAP) value. The manufactured model will enable the identification of R. pedestris in the field and can be an effective tool for insect forecasting in the early stage of pest outbreaks in crop production.

Special issue: Relationship between insect vectors and plant pathogens

Special issue: Relationship between insect vectors and plant pathogens

Symbiont coordinates stem cell proliferation, apoptosis, and morphogenesis of gut symbiotic organ in the stinkbug-Caballeronia symbiosis.

The bean bug Riptortus pedestris obtains a specific bacterial symbiont, Caballeronia insecticola (Burkholderia insecticola), from the environmental soil and harbors it in the posterior midgut region that is composed of hundreds of crypts. While newly hatched aposymbiotic insects possess primordial midgut crypts with little or no lumen, colonization of C. insecticola triggers swift development of the symbiotic organ, forming enlarged and opened crypts, and the symbiont subsequently fills the luminal cavities of those mature crypts. The cellular processes of crypt development triggered by C. insecticola colonization are poorly understood. Here we identified a fundamental mechanism of the symbiont-mediated midgut development by investigating cell cycles of intestinal epithelial cells. Intestinal stem cells of the bean bug are located and proliferate at the crypt base. Differentiated enterocytes migrate upward along the epithelial cell layer of the crypt as the midgut develops, induction of apoptosis in enterocytes primarily occurred on the tip side of the crypts, and apoptotic cells then eventually were shed from the crypts into the hemolymph. The proliferation rate of the stem cells at the base of the crypts was low while a high apoptotic rate was observed at the crypt tip in aposymbiotic insects, resulting in undeveloped short crypts. On the contrary, the gut-colonizing C. insecticola promoted the proliferation of the stem cells at the base of crypts and simultaneously inhibited apoptosis at the tip of crypts, resulting in a net growth of the crypts and the generation of a crypt lumen that becomes colonized by the bacterial symbiont. These results demonstrated that the Caballeronia symbiont colonization induces the development of the midgut crypts via finely regulating the enterocyte cell cycles, enabling it to stably and abundantly colonize the generated spacious crypts of the bean bug host.

Hadronotus pubescens (Motschoulsky) (Hymenoptera, Scelionidae): Redescription, biological attributes, and parasitism on eggs of Riptortus pedestris (Fab.) (Hemiptera, Alydidae)

Riptortus pedestris (Fab.) (Hemiptera, Alydidae) is one of the most damaging insects of leguminous crops in Eastern Asia but has only recently emerged as a pest in Bangladesh. Eggs, nymphs and adults of R. pedestris are here reported from mung bean (Vigna radiata (L.)) fields in Bangladesh. Two parasitoid species were reared from field-collected eggs of R. pedestris, the solitary Hadronotus pubescens (Motschoulsky) (Hymenoptera, Scelionidae) and a gregarious species of Ooencyrtus Ashmead (Hymenoptera, Encyrtidae). Here we redescribe H. pubescens, treat H. hogenakalensis (Sharma) as a junior synonym, and report aspects of its biology that were investigated under laboratory conditions. The number of eggs parasitized by H. pubescens was constant with eggs up to 48 hours in age, decreasing by 14% for 96 hour-old eggs. As host egg age increased, the parasitoid mean development time increased and the longevity of the parasitoids decreased.

Transcriptional profiling reveals a critical role of GmFT2a in soybean staygreen syndrome caused by the pest Riptortus pedestris.

Soybean staygreen syndrome, characterized by delayed leaf and stem senescence, abnormal pods, and aborted seeds, has recently become a serious and prominent problem in soybean production. Although the pest Riptortus pedestris has received increasing attention as the possible cause of staygreen syndrome, the mechanism remains unknown. Here, we clarify that direct feeding by R.pedestris, not transmission of a pathogen by this pest, is the primary cause of typical soybean staygreen syndrome and that critical feeding damage occurs at the early pod stage. Transcriptome profiling of soybean indicated that many signal transduction pathways, including photoperiod, hormone, defense response, and photosynthesis, respond to R.pedestris infestation. Importantly, we discovered that members of the FLOWERING LOCUS T (FT) gene family were suppressed by R.pedestris infestation, and overexpression of floral inducer GmFT2a attenuates staygreen symptoms by mediating soybean defense response and photosynthesis. Together, our findings systematically illustrate the association between pest infestation and soybean staygreen syndrome and provide the basis for establishing a targeted soybean pest prevention and control system.

Characterization of Burkholderia bacteria clade compositions in soil and Riptortus pedestris (Hemiptera: Alydidae) in South Korea

• Riptortus pedestris acquires bacterial symbiont, Burkholderia spp., from soil. • We investigated Burkholderia clade compositions using PCR analysis in South Korea. • Triple clade (SBE+ PBE+BCC) of Burkholderia was dominantly detected in soil. • Riptortus pedestris mainly acquired single BCC clade when reared on the soil samples. • From wild R. pedestris collected, the insects mainly harbored single BCC clade. Riptortus pedestris (Hemiptera: Alydidae) is known to acquire the genus Burkholderia , symbiotic bacteria, from soil. Therefore, symbiont acquisition of R. pedestris would be directly affected by bacterial diversity in soil. Soil typically harbors diverse microbes including different Burkholderia clades such as SBE (stinkbug-associated beneficial and environmental), PBE (plant-associated beneficial and environmental), and BCC ( Burkholderia cepacia and complex). Nevertheless, little is known about Burkholderia acquisition patterns of R. pedestris in nature, especially in the context of bacteria clade compositions in soil. Therefore, based on diagnostic PCR analysis, we investigated Burkholderia clade compositions in field-collected soil itself and R. pedestris when the insects were provided with the soil. Also, wild R. pedestris were surveyed to characterize their Burkholderia compositions. First, 88.44% of soil samples were detected with the genus Burkholderia , and triple clade (SBE+PBE+BCC) was most frequently detected. Second, R. pedestris nymphs readily acquired Burkholderia bacteria from field-collected soil where 91.25% of the reared insects harbored the bacteria in their midguts. In contrast to soil, the detection of single BCC clade was the most dominant among the three identified Burkholderia clades. Third, from wild R. pedestris , 80.62% of the insects were found to harbor the genus Burkholderia , and single BCC clade was most frequently detected. Finally, 29.13% and 47.06% of the reared and wild R. pedestris were detected with unidentified Burkholderia clade, which does not belong to any of the three identified clades. Our findings provide baseline information to better understand ecological associations between R. pedestris and Burkholderia bacteria in different clades.

Burkholderia gut symbiont induces insect host fecundity by modulating Kr-h1 gene expression.

Full-length cDNAs of the Broad-Complex (BR-C) from Riptortus pedestris were cloned. Moreover, Kr-h1 and BR-C expression levels in apo-symbiotic and symbiotic host insects were compared to verify whether they are modulated by Burkholderia gut symbionts. Interestingly, Kr-h1 expression level was significantly increased in symbiotic females. To determine how Kr-h1 affects fecundity in insects, the biosynthesis of two reproduction-associated proteins, hexamerin-α and vitellogenin, was investigated in R. pedestris females. Hexamerin-α and vitellogenin expression at the transcriptional and translational levels decreased in Kr-h1-suppressed symbiotic females, subsequently reduced egg production. These results suggest that Burkholderia gut symbiont modulates Kr-h1 expression to enhance ovarian development and egg production of R. pedestris by increasing the biosynthesis of the two proteins.

The Characterization of Three Novel Insect-Specific Viruses Discovered in the Bean Bug, Riptortus pedestris.

Insect-specific virus (ISV) is one of the most promising agents for the biological control of insects, which is abundantly distributed in hematophagous insects. However, few ISVs have been reported in Riptortus pedestris (Fabricius), one of the major pests threatening soybeans and causing great losses in yield and quality. In this work, field Riptortus pedestris was collected from six soybean-producing regions in China, and their virome was analyzed with the metatranscriptomic approach. Altogether, seven new insect RNA viruses were identified, three of which had complete RNA-dependent RNA polymerase (RdRp) and nearly full-length genome sequences, which were named Riptortus pedestris alphadrosrha-like virus 1 (RpALv1), Riptortus pedestris alphadrosrha-like virus 2 (RpALv2) and Riptortus pedestris almendra-like virus (RiALv). The three identified novel ISVs belonged to the family Rhabdoviridae, and phylogenetic tree analysis indicated that they were clustered into new distinct clades. Interestingly, the analysis of virus-derived small-interfering RNAs (vsiRNAs) indicated that only RiALv-derived siRNAs exhibited 22 nt length preference, whereas no clear 21 or 22 nt peaks were observed for RpALv1 and RpALv2, suggesting the complexity of siRNA-based antiviral immunity in R. pedestris. In conclusion, this study contributes to a better understanding of the microenvironment in R. pedestris and provides viral information for the development of potential soybean insect-specific biocontrol agents.

Screening and evaluation of entomopathogenic fungi against the bean bug, Riptortus pedestris using multiple tools

AbstractThe bean bug Riptortus pedestris causes major damage to soybean and has become a serious problem due to its high resistance to insecticides. To address this problem, this study aimed to select the most effective entomopathogenic fungi for the control of R. pedestris. Selection was performed through the evaluation of virulence, thermotolerance, and UV‐B tolerance of fungal isolates against R. pedestris and conidial production on grain medium. The pathogenicity of 30 Beauveria bassiana isolates and 30 Metarhizium anisopliae isolates was evaluated, and pathogenicity was confirmed in all but 3 isolates of B. bassiana. Among the 57 isolates, 11 exhibited high virulence. Of these 11 isolates, B. bassiana JB13G1W1 and M. anisopliae CB6T1G1 and JB12R3G1 showed the highest virulence of more than 80%. The conidial production of all fungal isolates in millet grain medium was good, ranging from 108 to 109 conidia/g. The UV‐B tolerance and thermotolerance of the 57 isolates varied according to the isolate, and those of M. anisopliae isolates were slightly higher than those of B. bassiana. As a result of comparative evaluation of the virulence, conidial production, thermotolerance, and UV‐B tolerance of fungal isolates, B. bassiana JB13G1W1 and M. anisopliae CB6T1G1 and JB12R3G1 were selected as excellent fungal isolates for the control of R. pedestris. Among them, B. bassiana JB13G1W1 showed the highest virulence, with LC50 and LT50 values of 7.86 × 105 conidia/mL and 5.24 days at 108 conidia/mL, respectively. The most effective growth stage to control R. pedestris with B. bassiana JB13G1W1 was 2nd‐instar nymphs.

Polyandrous females but not monogamous females vary in reproductive ageing patterns in the bean bug Riptortus pedestris

BackgroundIn general, reproductive performance exhibits nonlinear changes with age. Specifically, reproductive performance increases early in life, reaches a peak, and then declines later in life. Reproductive ageing patterns can also differ among individuals if they are influenced by individual-specific strategies of resource allocation between early-life reproduction and maintenance. In addition, the social environment, such as the number of available mates, can influence individual-specific resource allocation strategies and consequently alter the extent of individual differences in reproductive ageing patterns. That is, females that interact with more partners are expected to vary their copulation frequency, adopt a more flexible reproductive strategy and exhibit greater individual differences in reproductive ageing patterns.MethodsIn this study, we evaluated the effect of mating with multiple males on both group- and individual-level reproductive ageing patterns in females of the bean bug Riptortus pedestris by ensuring that females experienced monogamous (one female with one male) or polyandrous conditions (one female with two males).ResultsWe found that group-level reproductive ageing patterns did not differ between monogamy-treatment and polyandry-treatment females. However, polyandry-treatment females exhibited among-individual variation in reproductive ageing patterns, while monogamy-treatment females did not.ConclusionOur findings provide the first empirical evidence regarding the influence of the social environment on individual variation in reproductive ageing patterns. We further suggest that the number of potential mates influences group- and individual-level reproductive ageing patterns, depending on which sex controls mating. We encourage future studies to consider interactions between species-specific mating systems and the social environment when evaluating group- and individual-level reproductive ageing patterns.

Functional Analysis of a Juvenile Hormone Inducible Transcription Factor, Krüppel homolog 1, in the Bean Bug, Riptortus pedestris.

Juvenile hormone (JH) has two major functions in insects, i.e., suppression of metamorphosis in the larval or nymphal stage and promotion of reproduction in the adult stage. Krüppel homolog 1 (Kr-h1), a C2H2 zinc-finger type transcription factor, is reported to act downstream of the JH receptor complex. In the present study, the function of Kr-h1 was examined in adults and nymphs of Riptortus pedestris by RNA interference (RNAi). After injection of adults with dsRNA of Kr-h1, the expression level of Kr-h1 was significantly decreased in the abdomen. Kr-h1 dsRNA-injection resulted in a lower proportion of individuals with developed ovaries, but the difference was not statistically significant. The transcript levels of cyanoprotein-α and vitellogenin-1, which are JH-inducible genes encoding yolk proteins, were not affected in the abdomen by Kr-h1 knockdown. Kr-h1 dsRNA-injection was effective for suppression of Kr-h1 expression in nymphs. Some Kr-h1 dsRNA-injected fifth (final) instar nymphs had morphological defects in the wing bud. Moreover, they had several adult morphological features, including ocelli in the head, connexivum in the abdomen, coloring of the dorsal abdomen, and genitals. The nymphs possessing adult features did not emerge as adults during 1 month. These results demonstrated that Kr-h1 is necessary for maintaining nymphal characters in R. pedestris. The function of Kr-h1 in ovarian development remains unclear in R. pedestris.

Impact of Temperature on Age–Stage, Two-Sex Life Table Analysis of a Chinese Population of Bean Bug, Riptortus pedestris (Hemiptera: Alydidae)

Riptortus pedestris Fabricius (Hemiptera: Alydidae) is an economically important pest of soybean that causes serious damage in the Huang-Huai-Hai region of China, which is the main soybean-producing area. However, the basic biological and ecological attributes of the Chinese populations of R. pedestris are still unknown. Temperature is a critical factor influencing insect population dynamics; therefore, understanding the effects of temperature on individual development and population growth of R. pedestris is helpful to forecast the pest’s phenology as part of its control. In this study, we determined age–stage, two-sex life table parameters of R. pedestris at five constant temperatures (19, 22, 25, 28, and 31 °C) under laboratory conditions. The results demonstrated that the developmental period, preadult time, adult longevity, adult preoviposition period (APOP), and total preoviposition period (TPOP) were all significantly shortened when the temperature increased from 19 to 31 °C. The developmental time from egg to adult was 55.00 d at 19 °C but decreased to 24.43 d at 31 °C. The developmental threshold temperatures of the egg, nymph, female adult, male adult, and total generation (from egg to adult death) were 13.31, 10.91, 16.21, 17.13, and 13.44 °C, respectively, and the corresponding stage-specific day degree totals were 95.91, 260.51, 114.22, 147.35, and 582.04. The net reproductive rate (R0 = 2.36 offspring), fecundity (58.00 ± 28.59 eggs), and population trend index (I) (7.89) were found to be optimal at 25 °C. These results could be useful for predicting the population dynamics of R. pedestris and improving the management of this pest in field crops.

Clock gene-dependent glutamate dynamics in the bean bug brain regulate photoperiodic reproduction.

Animals adequately modulate their physiological status and behavior according to the season. Many animals sense photoperiod for seasonal adaptation, and the circadian clock is suggested to play an essential role in photoperiodic time measurement. However, circadian clock-driven neural signals in the brain that convey photoperiodic information remain unclear. Here, we focused on brain extracellular dynamics of a classical neurotransmitter glutamate, which is widely used for brain neurotransmission, and analyzed its involvement in photoperiodic responses using the bean bug Riptortus pedestris that shows clear photoperiodism in reproduction. Extracellular glutamate levels in the whole brain were significantly higher under short-day conditions, which cause a reproductive diapause, than those under long-day conditions. The photoperiodic change in glutamate levels was clearly abolished by knockdown of the clock gene period. We also demonstrated that genetic modulation of glutamate dynamics by knockdown of glutamate-metabolizing enzyme genes, glutamate oxaloacetate transaminase (got) and glutamine synthetase (gs), attenuated photoperiodic responses in reproduction. Further, we investigated glutamate-mediated photoperiodic modulations at a cellular level, focusing on the pars intercerebralis (PI) neurons that photoperiodically change their neural activity and promote oviposition. Electrophysiological analyses showed that L-Glutamate acts as an inhibitory signal to PI neurons via glutamate-gated chloride channel (GluCl). Additionally, combination of electrophysiology and genetics revealed that knockdown of got, gs, and glucl disrupted cellular photoperiodic responses of the PI neurons, in addition to reproductive phenotypes. Our results reveal that the extracellular glutamate dynamics are photoperiodically regulated depending on the clock gene and play an essential role in the photoperiodic control of reproduction via inhibitory pathways.