Pupal Stage Research Articles

Rhodamine-B for the mark, release, and recapture experiments in gamma-irradiated male Aedes aegypti (Diptera: Culicidae): Persistence, dispersal, and its effect on survival.

Rhodamine-B (Rh-B) marking shows a great potential for use in mark-release-recapture (MRR) studies for rear-and-release mosquito control strategies, including the radiation-based sterile insect technique. However, its applicability and evaluation in body-stain-irradiated males of Aedes aegypti have received little attention. The present study evaluated the use of Rh-B to mark gamma-irradiated male A. aegypti. Male A. aegypti were irradiated at the pupal stage at a dose of 70 Gy. After emergence, males were fed 0.1, 0.2, 0.3, or 0.4% Rh-B in 10% glucose solution for 4 days. Groups of unirradiated males that received the same feeding treatments were used as control groups. We evaluated the persistence of Rh-B and the longevity of males after Rh-B feeding. Furthermore, the use of Rh-B in irradiated A. aegypti for MRR experiments was evaluated at an urban site. No difference was observed in the Rh-B persistence among all concentrations at the 24-h postmarking period ranging from 91.25 ± 1.61% to 96.25 ± 1.61% and from 90.00 ± 2.28% to 93.13 ± 2.77% for the unirradiated and irradiated groups, respectively. Rh-B persistence significantly decreased over time, and persistence was significantly longer with increased concentrations in both the unirradiated and irradiated groups. Longevity was considerably decreased by Rh-B feeding and irradiation. However, no significant difference in longevity was found among males fed various concentrations of Rh-B. Through MRR experiments, irradiated-Rh-B marked males were mostly detected within a radius of 20 m and 40 m from the center-release point. The mean distance traveled of the released males from the three MRR events was calculated to be 42.6 m. This study confirms that Rh-B body marking through sugar feeding is applicable for irradiated male A. aegypti, with only a slight effect on longevity. Furthermore, considering the significant reduction in persistence over time, further study is needed to assess the impact of this reduction on the calculation of field biological parameters resulting from MRR experiments.

SlCarE054 in Spodoptera litura (Lepidoptera: Noctuidae) showed direct metabolic activity to β-cypermethrin with stereoselectivity.

Carboxylesterases (CarEs) is an important detoxification enzyme system in phase Ⅰ participating in insecticides resistance. In our previous study, SlCarE054, a CarEs gene from lepidoptera class, was screened out to be upregulated in a pyrethroids and organophosphates resistant population. Its overexpression was verified in two field-collected populations of Spodoptera litura (Lepidoptera: Noctuidae) resistant to pyrethroids and organophosphates by qRT-PCR. Spatiotemporal expression results showed that SlCarE054 was highly expressed in the pupae stage and the digestive tissue midgut. To further explore its role in pyrethroids and organophosphates resistance, its metabolism activity to insecticides was determined by UPLC. Its recombinant protein showed significant metabolism activity to cyhalothrin and fenvalerate, but not to phoxim or chlorpyrifos. The metabolic activity of SlCarE054 to β-cypermethrin showed stereoselectivity, with higher metabolic activity to θ-cypermethrin than the enantiomer α-cypermethrin. The metabolite of β-cypermethrin was identified as 3-phenoxybenzaldehyde. Further modelling and docking analysis indicated that β-cypermethrin, cyhalothrin and fenvalerate could bind with the catalytic triad of the 3D structure of SlCarE054. The interaction of β-cypermethrin with SlCarE054 also showed the lowest binding energy. Our work provides evidence that SlCarE054 play roles in β-cypermethrin resistance in S. litura.

The neonicotinoid acetamiprid reduces larval and adult survival in honeybees (Apis mellifera) and interacts with a fungicide mixture

Plant protection products (PPPs), which are frequently used in agriculture, can be major stressors for honeybees. They have been found abundantly in the beehive, particularly in pollen. Few studies have analysed effects on honeybee larvae, and little is known about effects of insecticide-fungicide-mixtures, although this is a highly realistic exposure scenario. We asked whether the combination of a frequently used insecticide and fungicides would affect developing bees. Honeybee larvae (Apis mellifera carnica) were reared in vitro on larval diets containing different PPPs at two concentrations, derived from residues found in pollen. We used the neonicotinoid acetamiprid, the combined fungicides boscalid/dimoxystrobin and the mixture of all three substances. Mortality was assessed at larval, pupal, and adult stages, and the size and weight of newly emerged bees were measured. The insecticide treatment in higher concentrations significantly reduced larval and adult survival. Interestingly, survival was not affected by the high concentrated insecticide-fungicides-mixture. However, negative synergistic effects on adult survival were caused by the low concentrated insecticide-fungicides-mixture, which had no effect when applied alone. The lower concentrated combined fungicides led to significantly lighter adult bees, although the survival was unaffected. Our results suggest that environmental relevant concentrations can be harmful to honeybees. To fully understand the interaction of different PPPs, more combinations and concentrations should be studied in social and solitary bees with possibly different sensitivities.

Effect of ultraviolet on the environmental adaptability of Megalurothrips usitatus (Thysanoptera: Thripidae).

Megalurothrips usitatus Bagnall, an important pest of bean plants, is primarily managed with synthetic insecticides. M. usitatus has developed considerable resistance to various insecticides in multiple cowpea-growing areas in Hainan Province, China, posing challenges to its control in the field. Light control technology is a potentially effective physical control method for M. usitatus. The vision of thrips is highly sensitive to UV light, whereas other biological characteristics remain unknown. Therefore, this study evaluated the effects of ultraviolet light on the biological characteristics of M. usitatus. Results showed that the egg, larval, and pupal stages of M. usitatus were significantly shortened, and the emergence rate (79.59%) and adult survival rate (77.95%) were reduced under a devoid of UV light environment (UV-), compared with the full-spectrum light (control treatment group, CK) (p < 0.05). However, the single spawning quantity and total amount of spawning were significantly higher, and the sex ratio (57%) was the highest under UV- (p < 0.05). Single UV light (UV+) only affected the pupation rate. Also, the antioxidant enzymes, polyphenol oxidase, superoxide dismutase (SOD), and peroxidase activities were significantly and negatively correlated with the progression of generations under UV-, whereas catalase and SOD activities were significantly and positively correlated with the progression of generations under UV+. The UV- light conditions significantly interfered with the behavior selection of M. usitatus. The results of this study showed that the adaptability of M. usitatus populations would be greatly reduced in the absence of ultraviolet light, providing a theoretical basis for the control of M. usitatus populations.

Impact of Maize Nutrient Composition on the Developmental Defects of Spodoptera frugiperda

Spodoptera frugiperda (J.E. Smith) is a crucial agricultural pest owing to its global impact on &gt;300 crops. Among these, the corn strain of S. frugiperda causes significant damage to maize (Zea mays L.). However, limited research exists on the influence of maize nutrients on the metamorphosis of S. frugiperda and the underlying mechanisms. In this study, the effects of different growth stages of maize leaves, namely, tender leaves (tender) and mature leaves (mature), on various aspects of larval development, including body weight, body length, developmental age, pupation rate, and eclosion rate, were investigated. Additionally, we measured the levels of 20-hydroxyecdysone (20E) and three types of juvenile hormone (JH; i.e., JH I–III) in S. frugiperda larvae fed on tender or mature. The results revealed that larvae fed on Tender exhibited significantly prolonged instar duration, reduced body weight and length, and decreased pupation and eclosion rates, with the emergence of abnormal adults. Analysis of nutritional components in maize leaves revealed significantly higher levels of amino acids, soluble sugars, and sterols in mature than in tender. Hormone analysis in S. frugiperda larvae revealed higher 20E titers in individuals feeding on mature during prepupal and pupal stages. We demonstrated the crucial role of sterols in regulating the level of 20E and pupation rate of S. frugiperda. Based on these findings, we propose that isoleucine, arginine, glutamic acid, sucrose, campesterol, and β-sitosterol serve as key nutrients influencing the development of S. frugiperda. Moreover, β-sitosterol is a significant factor influencing the interaction between maize leaves and S. frugiperda. Our research results provide a reference for the control strategy of S. frugiperda based on breeding insect-resistant varieties by altering host nutrition.

Structural changes shaping the Drosophila ellipsoid body ER-neurons during development and aging

The ellipsoid body (EB) of the insect brain performs pivotal functions in controlling navigation. Input and output of the EB is provided by multiple classes of R-neurons (now referred to as ER-neurons) and columnar neurons which interact with each other in a stereotypical and spatially highly ordered manner. The developmental mechanisms that control the connectivity and topography of EB neurons are largely unknown. One indispensable prerequisite to unravel these mechanisms is to document in detail the sequence of events that shape EB neurons during their development. In this study, we analyzed the development of the Drosophila EB. In addition to globally following the ER-neuron and columnar neuron (sub)classes in the spatial context of their changing environment we performed a single cell analysis using the multi-color flip out (MCFO) system to analyze the developmental trajectory of ER-neurons at different pupal stages, young adults (4d) and aged adults (∼60d). We show that the EB develops as a merger of two distinct elements, a posterior and anterior EB primordium (prEBp and prEBa, respectively. ER-neurons belonging to different subclasses form growth cones and filopodia that associate with the prEBp and prEBa in a pattern that, from early pupal stages onward, foreshadows their mature structure. Filopodia of all ER-subclasses are initially much longer than the dendritic and terminal axonal branches they give rise to, and are pruned back during late pupal stages. Interestingly, extraneous branches, particularly significant in the dendritic domain, are a hallmark of ER-neuron structure in aged brains. Aging is also associated with a decline in synaptic connectivity from columnar neurons, as well as upregulation of presynaptic protein (Brp) in ER-neurons. Our findings advance the EB (and ER-neurons) as a favorable system to visualize and quantify the development and age-related decline of a complex neuronal circuitry.

Review: Nutritional Needs of Honeybees and Legislation on Apiculture By-Products in Animal Nutrition.

Honeybees are some of the smallest farmed animals, and apiculture by-products, e.g., honey, beeswax, propolis, royal jelly, and pollen contribute to animal nutrition. For the effective production of these by-products, the optimal development and nutrient supply of the honeybee is required. Beginning with the development of the mouth and anal pores on the second day of embryonic development, the digestive tract differentiates into the mouth and fore-, mid-, and hindgut during the pupal stage. The various glands within the oral cavity are particularly important, secreting enzymes and substances that are crucial for digestion and hive nutrition, e.g., invertase and royal jelly. Honeybees rely on a specialized caste system, with worker bees collecting nectar, pollen, water, and resin for the nutrition of the entire hive. Macronutrients, including proteins, carbohydrates, and lipids, obtained primarily from pollen and nectar, are essential for the growth and development of larvae and the overall health of the colony. Inadequate nutrient intake can lead to detrimental effects on larval development, prompting cannibalism within the hive. Apiculture by-products possess unique nutritional and therapeutic properties, leading to a growing interest in the use of honey, beeswax, propolis, and pollen as a feed additive. In recent years, the use of apicultural by-products in animal nutrition has been primarily limited to in vivo studies, which have demonstrated various positive impacts on the performance of farm animals. Honey, beeswax, propolis, royal jelly, and pollen are listed feed stuffs according to Regulation (EC) No. 68/2013. However, for animal nutrition there is not any specific legal definition for these products and no legal requirements regarding their ingredients as given for honey or beeswax in European food law.

Comparative Population Biology and Related Gene Expression in the Beta-Cypermethrin-Resistant Strains of Bactrocera dorsalis (Hendel).

Diptera and Lepidoptera species have the highest levels of insecticide resistance, and the mechanism of drug resistance has been studied in detoxification metabolism genes such as P450, GST, EST, and ABC. Since Bactrocera dorsalis are resistant to a variety of chemicals, the pattern and mechanism of resistance in Bactrocera dorsalis have been investigated from a variety of aspects such as detoxification metabolism genes, detoxification enzymes, intestinal symbiotic bacteria, and synergists in the world. In this study, 51 species and 149 detoxification metabolism genes were annotated in the Suppression Subtractive Hybridization (SSH) library, and 12 candidate genes related to beta-cypermethrin resistance were screened and quantitatively expressed in this library. Two genes were found to be upregulated in the egg stage, three genes in the larval stage, one gene in the pupal stage, and five genes in the adult stage, and four genes were found to be upregulated in the midgut and the malacca ducts in the midgut. The expression of cyp6g1, cyp6a22, GST-Epsilon9, and Trypsin-4 genes was upregulated in resistant strains, with the most obvious upregulation occurring in the midgut and the Malpighian tubules. These results provide new insights into the study of pesticide resistance in quarantine insects.

An Internal Marking Method for Adult Spodoptera frugiperda Smith Using an Artificial Diet Containing Calco Oil Red N-1700.

As a migratory invasive pest, Spodoptera frugiperda (fall armyworm, FAW) has recently posed a serious threat to food security in newly invaded areas (especially in Africa and Asia). Understanding its migration (or dispersal) patterns in newly invaded areas is crucial for regional forecasting and management efforts. By screening an appropriate marking technique to conduct mark-release-recapture (MRR) experiments, the migration patterns of the FAW can be effectively studied. In this study, we added different concentrations of Calco Oil Red N-1700 (an oil-soluble marker) to a self-made artificial diet and assessed the rearing and marking efficacy. The results indicated that a concentration of 0.2% of Calco Oil Red N-1700 in the diet was optimal for marking adult FAWs. The biological indicators (e.g., developmental duration, reproductive parameters, and flight ability) of FAWs fed this diet were basically consistent with those of FAWs fed a normal diet, with a larval stage of 15.46 days, a pupal stage of 9.81 days, a pupal mass of 278.18 mg, an adult longevity of 15.41 days, and an egg deposition count of 1503.51. Meanwhile, the flight distance, duration, and velocity were 24.91 km, 7.16 h, and 3.40 km/h, respectively (12 h tethered-flight tests), without difference with the control. Females and males exhibited distinctive marking colors (red or pink) that persisted for at least 5 and 9 days, respectively. This study developed an economically effective internal marking method for the adult FAW, laying the foundation for conducting MRR experiments. This will help clarify the migration behavior and routes of the FAW, providing a scientific basis for formulating effective pest management strategies.

The genus Acerocephala and observations of the life history of Acerocephala hanuuanamu sp. nov. (Hymenoptera, Cerocephalidae) and its bark beetle host on the island of O‘ahu, Hawai‘i

The behavior of Acerocephala hanuuanamusp. nov. found parasitizing Cryphalus brasiliensis under the surface of wood in Ficus microcarpa trees on the island of O‘ahu in Hawai‘i is deduced from observations in naturally occuring branches, and from direct observation using specially designed “phloem sandwich” observation chambers which consist of tree bark peeled through the phloem layer from freshly cut branches and sandwiched between a sheet of aluminum and a sheet of plexiglass. Cryphalus brasiliensis beetles, found living in large numbers in the phloem tissues in F. microcarpa trees, were collected and placed into these chambers. They tunneled into the wood and reproduced, producing an active colony of all life stages. Acerocephala hanuuanamu wasps were then placed into the system and their behavior observed. Typical behavior, aspects of which were recorded in video and still images, was as follows. A female A. hanuuanamu enters the tunnels of the bark beetles. She digs through the debris in the tunnels in a search for larvae and pupae. Cryphalus brasiliensis prepupae construct a hard pupal chamber around themselves before they pupate, and upon encountering a larva in the tunnels or the exterior of a pupal chamber with a pupa inside, she adeptly turns around in the tunnel to sting it, either inserting her ovipositor directly into the larva or by laboriously pushing it through the hard shell of the pupal chamber. When finished stinging, she withdraws her ovipositor slowly and carefully, often extracting tissue from the beetle immature as a sheath around her ovipositor. This structure remains projecting from the larva or pupa, and then the wasp turns around and host feeds through this structure, in the case of a pupa at a substantial distance through the wall of the pupal chamber. Oviposition occurs on larval stages and pupal stages. The egg hatches and the larva develops as an ectoparasitoid on the beetle. When finished feeding, it detaches and pupates in the tunnel. Mating behavior is also described. In addition to C. brasiliensis in F. microcarpa, A. hanuuanamu was also observed attacking Cryphalus mangiferae in mango (Mangifera indica) branches. Acerocephala ihulenasp. nov. was also found on O‘ahu parasitizing Eidophelus pacificus in hau (Hibiscus tiliaceus) branches, and is described. The genus Acerocephala is revised given the perspective resulting from these two new species and aspects of functional morphology observed in the behavioral studies, Acerocephala indicacomb. nov. is transferred to the genus, and a key to the genus is provided.

The combined effect of sublethal concentrations of insecticides and local entomopathogenic nematode isolates on larval and pupal stages of Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae)

The black cutworm (BCW), Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae), is an economically significant pest affecting numerous crops and turf grasses. Despite employing various methods, such as chemical and biological control, to mitigate this pest, the efficacy of these methods often fails to reach the desired level. Hence, the integration of diverse strategies is a key to sustainable management. In this study, we assessed the toxicity of four insecticides (indoxacarb, pyridalyl, lufenuron, and methoxyfenozide) and three local EPN isolates (two isolates of Heterorhabditis indica and one of Heterorhabditis bacteriophora) in targeting the fifth instar larval (L5) and pupal stages of BCW. After demonstrating their potential use in BCW control, we explored the combined use of sublethal concentrations of insecticides (LC15 or LC30) alongside the LC20 values of EPNs. The findings revealed that lufenuron and methoxyfenozide, when combined with EPNs, exhibited a synergistic effect on the L5 stage of BCW, whereas the same combination displayed antagonism on pupae. On the other hand, the combination of indoxacarb and pyridalyl with EPNs demonstrated a synergistic effect on the pupal stage of BCW. These results highlight the potential of employing a combination of insecticides and EPNs at sublethal concentrations to control BCW effectively. This approach shows promise for designing sustainable pest management strategies within the framework of integrated pest management (IPM).

Performance of Hermetia illucens reared on pure, mixed, and sequentially mixed organic wastes regionally available in central Argentina

Abstract Black soldier fly larvae (Hermetia illucens L.) are proficient decomposers of organic matter. We compared experiments with local substrates containing: domestic residues (DR), and brewery wastes (BW) in pure form, or mixed with in two different proportions (BW70 or BW30), and two sequential diets in which larvae were fed first DR and then BW (DR/BW) or vice versa (BW/DR). The effects of the diets on life cycle parameters for mass production of H. illucens larvae were evaluated using general linear mixed models, ANOVA and Kruskal-Wallis. The pure DR diet significantly prolonged the developmental period (47.8 days), although the pupal stage was shorter in larvae fed on this diet (13.8 days). Larval survival was similar between treatments (over 85%), but pupal survival tended to be higher in the mixed diets (over 96%). Larval weights differed significantly between treatments, with mixed diets promoting greater weight gains than pure and sequential diets. Larvae fed a DR/BW diet gained more weight than those fed a BW/DR diet. The highest mean larval weights (0.23 g) were observed in the mixed diet BW70, followed by the sequential diet DR/BW (0.22 g). Mixed diet BW70 also resulted in larger wings and bodies in both sexes. Sequential diet BW/DR and pure diet DR resulted in the smallest adult sizes. Sex ratios varied, favouring male emergence (1.8 M:0.2 F) in pure diet DR and in both sequential mixed diets (1.7 M:0.3 F and 1.3 M:0.7 F, respectively). This study highlights the importance of higher total protein plus carbohydrate concentrations in diets for optimal larval rearing, favouring mixed diets BW70. Carbohydrate-rich substrates should be provided earlier in the cycle to benefit larval and adult performance. Locally available waste vegetable or fruit peels, brewery waste, and spent yerba mate proved to be viable substrates for larval rearing, providing economic and environmental benefits for H. illucens rearing.

Timing-dependent effects of elevated temperature on reproductive traits in the European corn borer moth.

Elevated temperature often has life stage-specific effects on ectotherms because thermal tolerance varies throughout ontogeny. Impacts of elevated temperature may extend beyond the exposed life stage, if developmental plasticity causes early exposure to carry over, or if exposure at multiple life stages cumulatively produces effects. Reproductive traits may be sensitive to different thermal environments experienced during development, but such effects have not been comprehensively measured in Lepidoptera. In this study, we investigate how elevated temperature at different life stages alters reproduction in the European corn borer moth, Ostrinia nubilalis. We tested effects of exposure to elevated temperature (28°C) separately or additively during larval, pupal, and adult life stages compared to control temperatures (23°C). We found that exposure to elevated pupal and adult temperature decreased the number of egg clusters produced, but exposure limited to a single stage did not significantly impact reproductive output. Furthermore, elevated temperature during the pupal stage led to a faster transition to the adult stage and elevated larval temperature altered synchrony of adult eclosion, either by itself or combined with pupal temperature exposure. These results suggest that exposure to elevated temperature during development alters reproduction in corn borers in multiple ways, including through carry-over and additive effects. Additive effects of temperature across life stages are thought to be less common than stage-specific or carry-over effects, but our results suggest thermal environments experienced at all life stages need to be considered when predicting reproductive responses of insects to heatwaves.

Seasonal life cycle of Pintomyia longiflocosa (Diptera: Psychodidae) under semi-controlled conditions in the Huila state, Colombia.

Pintomyia longiflocosa is considered a vector of etiological agents of cutaneous leishmaniasis. The objective of this research was to establish the parameters of the life cycle of Pi. longiflocosa in semi-controlled conditions in a rural area of the Campoalegre municipality, Huila, Colombia. The life cycle of individuals of Pi. longiflocosa, obtained from two cohorts of collected, fed, and individualized females, was monitored during two different times of the year (between February and August 2020 and between July 2020 and January 2021, respectively). Determining parameters associated with the fertility and fecundity, time and attributes of development and survival, and its association with abiotic variables. The average duration of Cycle 1 (C1) and Cycle 2 (C2) was 134.9 and 148.78 days, respectively. The gonotrophic cycle of parental females presented significant differences (p-value <0.05) between C1 and C2 (8.47 and 11.42 days) as well as between fecundity and fertility parameters. The number of days it takes the development of the immature stages between the two cycles studied, also showed significant differences in the larvae II (15.21 and 22.23), larvae III (11.93 and 17.56), and pupae (24.48 and 22.9) stages. During C1, the survival rate was higher and consistent with the productivity of adult individuals (F1), compared to C2. Fecundity and fertility values were significantly higher in C2. Finally, a significant correlation between the number of individuals and temperature was evidenced in C1 while, for C2, there was a negative correlation with precipitation. Significant differences were found in several biological and reproductive parameters between the two cycles monitored. The parameters of the life cycle of Pi. longiflocosa in its natural habitat would be influenced by environmental factors related to the annual seasonality in the sub-Andean rural area, conditioning the temporal distribution of this species and, consequently, the possible transmission of causative agents of cutaneous leishmaniasis.

Can the Necrophagous Blow Fly Calliphora vicina (Diptera: Calliphoridae) Be Reared on Plant-Based Meal?

The use of the blow fly Calliphora vicina as a potential pollination species to augment the current reliance on honeybees (Apis mellifera) in Australian horticulture requires knowledge of how best to mass-rear this fly species. Calliphora vicina lays eggs onto carrion soon after death, and the resultant larvae that hatch are necrophagous and feed on the decomposing tissues of the dead animal. Newly hatched larvae of this fly were provided with plant-based meals (soya bean and canola) and compared with larvae provided with livestock-derived meatmeal to determine if plant-based meal could be used to mass-rear this blow fly species. Both soya bean and canola meal media did not support larval survival through to adult emergence. The addition of only 10% whole egg powder to the plant-based meals enabled survival to eclosion of 39% and 13% on soya bean and canola-based media, respectively, compared with 76% on livestock-based meatmeal with 10% whole egg powder. Larvae fed a diet of livestock-based meatmeal with 10% whole dried egg powder had the fastest development to the pupal stage, the highest pupation rate, the heaviest pupae, and the highest subsequent adult eclosion. This study concluded that the use of plant-based meals as a diet for the mass-rearing of the blow fly C. vicina was not a viable option.

Establishing a Male-Positive Genetic Sexing Strain in the Asian Malaria Vector Anopheles stephensi.

Genetic biocontrol interventions targeting mosquito-borne diseases require the release of male mosquitoes exclusively, as only females consume blood and transmit human pathogens. This reduces the risk of spreading pathogens while enabling effective population control. Robust sex sorting methods to enable early larval sorting in mosquitoes need to be developed to allow for scalable sex sorting for genetic biocontrol interventions. This study applies the SEPARATOR (Sexing Element Produced by Alternative RNA-splicing of A Transgenic Observable Reporter) system, previously developed for Aedes aegypti, to the Asian malaria vector Anopheles stephensi. We hypothesized that the intron from the doublesex gene in Anopheles gambiae would function in An. stephensi due to evolutionary conservation. Our results confirm that the splicing module from An. gambiae operates effectively in An. stephensi, demonstrating evolutionary conservation in sex-specific splicing events between these species. This system enables reliable positive male selection from first instar larval to pupal stages. RT-PCR analysis demonstrates that male-specific EGFP expression is dependent on doublesex sex-specific splicing events. The SEPARATOR system's independence from sex-chromosome linkage confers resistance to meiotic recombination and chromosomal rearrangements. This approach may facilitate the mass release of males, and the cross-species portability of SEPARATOR establishes it as a valuable tool for genetic biocontrol interventions across various pest species.

A comparative study of fat body at different developmental stages in Bombyx mori (L.): Histological and ultrastructural aspects

AbstractThe fat body, originating from mesodermal tissue of coelomic cavities during embryogenesis, can be found throughout the insect's body. In some insects, the fat body has two types (peripheral and perivisceral fat bodies) according to its location. In the fat body, trophocytes are known to be the basic cell type. Other fat body cells differentiating from the trophocytes are urocytes, mycetocytes, chromatocytes and haemoglobin cells. The study aimed to demonstrate the general structure and cell types of the fat body during the postembryonic developmental stages of Bombyx mori (L.). With Haematoxylin–Eosin staining, there is a single type of fat body throughout the larval stages but during Pp and P0 stages it consists of perivisceral and peripheral fat bodies. We also observed that the compact structure of the fat body at the beginning of the larval stage began to loosen as it transformed into the pupal stage. In the adult stage, it was reorganized during the metamorphosis and it didn't show both fat bodies distinction. At this stage, the fat body appeared in a loose, lace‐like structure. We determined that B. mori fat body only consists of trophocytes and also among them; the fat body associated cells, oenocytes are detected.

The Lethal Toxic Effect of Alcoholic Extract of Punica granatum Plant on Stages of The Southern Cowpea beetle Callosobruchus maculatus (Coleoptera: Bruchidae)

This study was conducted to find out the toxic effect of the southern cowpea weevil Callosobruchus maculatus stages when its food was treated with alcoholic extracts of Punica granatum peels. The study showed that the lowest percentage of hatching was 61.26% in the treatment of chickpea seeds with a concentration of 15% of P. granatum peel extract. The treatments had a clear effect on the harmful larval stage , and the highest significant percentage of death was 81.4% in cowpea seeds treated with a concentration of 25%, and all results were significantly different from the control treatment. the mortality rate significantly increased in the pupa stage, and it increased treatment significantly to 46.66% in the concentration of 25%, compared to the, control which amounted to 0%. To find out the effect of the extracts used in the study on the first generation individuals, the egg retention coefficient was calculated.

Assessing black soldier fly pupation and survival in lunar regolith simulant: Implications for sustainable controlled habitats on the Moon

Bioregenerative life-support systems (BLSSs) will be crucial for extended space missions and extraterrestrial habitats. The black soldier fly, Hermetia illucens, is recognized for its efficient organic waste consumption, making it well-suited for closed environments like spacecraft. Our study assessed H. illucens adaptability to different substrates, including a lunar regolith simulant, pertinent to future lunar colonization. Remarkably resilient, H. illucens prepupal larvae successfully pupated in all tested substrates, but pupation timing varied, with no-substrate larvae pupating later. Pupal stage duration also differed, particularly with lunar regolith simulant and sand treatments resulting in longer durations. Substrate treatments significantly influenced the number of emerged adults, with lunar regolith simulant yielding more adults than the no-substrate treatment. Additionally, sand and wood shavings treatments produced more adults, highlighting H. illucens adaptability to various substrates, including lunar regolith. These findings are crucial for future BLSSs design. Additionally, H. illucens adaptability to lunar regolith provides insights into life's adaptability in space environments, guiding future experiments on celestial bodies. This study provides critical data on how different substrates, including lunar regolith simulants, influence H. illucens development and survival, advancing BLSSs and ecological science in both space and terrestrial contexts.

A simple PCR-based quick detection of the economically important oriental fruit fly, Bactrocera dorsalis (Hendel) from India.

The oriental fruit fly, Bactrocera dorsalis (Hendel), is a significant economic and quarantine pest due to its polyphagous nature. The accurate identification of B. dorsalis is challenging at the egg, maggot, and pupal stages, due to lack of distinct morphological characters and its similarity to other fruit flies. Adult identification requires specialized taxonomist. Existing identification methods are laborious, time consuming, and expensive. Rapid and precise identification is crucial for timely management. By analyzing the variations in the mitochondrial cytochrome oxidase-1 gene sequence (Insect barcoding gene), we developed a species-specific primer (SSP), DorFP1/DorRP1, for accurate identification of B. dorsalis. The optimal annealing temperature for the SSP was determined to be 66°C, with no cross-amplification or primer-dimer formation observed. The SSP was validated with B. dorsalis specimens from various locations in northern and eastern India and tested for cross-specificity with six other economically significant fruit fly species in India. The primer specificity was further confirmed by the analysis of critical threshold (Ct) value from a qPCR assay. Sensitivity analysis showed the primer could detect template DNA concentrations as low as 1 pg/µl, though sensitivity decreased at lower concentrations. Sequencing of the SSP-amplified product revealed over >99% similarity with existing B. dorsalis sequences in the NCBI GenBank. The developed SSP reliably identifies B. dorsalis across all developmental stages and sexes. This assay is expected to significantly impact pest identification, phytosanitary measures, and eradication programs for B. dorsalis.