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Plant-mediated RNA interference (RNAi) holds great promise for insect pest control, as plants can be transformed to produce double-stranded RNA (dsRNA) to selectively down-regulate insect genes essential for survival. For optimum potency, dsRNA can be produced in plant plastids, enabling the accumulation of unprocessed dsRNAs. However, the relative effectiveness of this strategy in inducing an RNAi response in insects using different feeding mechanisms is understudied. To investigate this, we first tested an in vitro-synthesized 189 bp dsRNA matching a highly conserved region of the v-ATPaseA gene from cotton mealybug (Phenacoccus solenopsis) on three insect species from two different orders that use leaf-chewing, lacerate-and-flush, or sap-sucking mechanisms to feed, and showed that the dsRNA significantly down-regulated the target gene. We then developed transplastomic Micro-tom tomato plants to produce the dsRNA in plant plastids and showed that the dsRNA is produced in leaf, flower, green fruit, red fruit, and roots, with the highest dsRNA levels found in the leaf. The plastid-produced dsRNA induced a significant gene down-regulation in insects using leaf-chewing and lacerate-and-flush feeding mechanisms, while sap-sucking insects were unaffected. Our results suggest that plastid-produced dsRNA can be used to control leaf-chewing and lacerate-and-flush feeding insects, but may not be useful for sap-sucking insects.

Background: Plastic pollution, particularly from polystyrene, has emerged as a serious environmental concern, prompting growing interest in its biodegradation. We investigated the potential of four chewing insects; mealworm (Tenebrio molitor), superworm (Zophobas morio), American cockroach (Periplaneta americana), and cricket (Gryllus bimaculatus) to biodegrade polystyrene paper (PSP). Methods: Using a randomized complete block design, four chewing insects were divided into two groups after 48-hour fasting with water. One group received PSP only; the other received PSP mixed with tapioca starch. Weight loss of PSP was recorded after 72 hours. Results: Z. morio demonstrated the highest degradation efficiency for PSP (92.10%), followed by P. americana (32.17%). When tapioca starch was added, Z. morio remained the highest effective (95.45%), followed by T. molitor (59.15%). Supplementing starch significantly enhanced degradation rates (P < 0.05). Fourier Transform Infrared Spectroscopy (FTIR) and Scanning Electron Microscopy (SEM) revealed signs of depolymerization, oxidation, and surface cracking. FTIR indicated new functional groups of carbonyl (C=O, 1650 cm⁻¹) and hydroxyl (O–H, 3200–3550 and 3584–3700 cm⁻¹). Conclusion: Certain chewing insects, especially Z. morio and T. molitor, possess strong potential for PSP degradation, likely aided by their gut microbiota. Notably, this study is the first to report PSP degradation by P. americana and G. bimaculatus. Further research is needed to explore the microbial mechanisms within insect guts that facilitate plastic biodegradation.

Abstract Transgenic Bt soybean plants have been developed to control insect pests, such as Anticarsia gemmatalis and Chrysodeixis includens. This objective has been achieved successfully; however, recently, some authors claimed that Bt soybean plants have been more susceptible than non-Bt soybean to Bemisia tabaci MEAM1. In addition, it is unknown whether Bt soybean plants infested by B. tabaci become less resistant to target pests. Therefore, this study aimed to evaluate: (i) whether the previous infestation with B. tabaci can compromise Bt and non-Bt soybean resistance to C. includens; (ii) the effects of B. tabaci infestations on Bt and non-Bt soybean plant growth; and (iii) whether B. tabaci feeding reduces contents of chlorophyll and carotenoids of soybean plants. Bt and non-Bt soybean plants pre-infested with B. tabaci showed no changes in resistance to C. includens. Bt soybean plants infested with B. tabaci showed a lower plant height than uninfested plants. Differently, non-Bt soybean plants exhibited no reduction in plant growth due to B. tabaci feeding. Bt soybean plants suffered a reduction in dry matter only under double infestation (B. tabaci and C. includens), while non-Bt soybean plants experienced reduction in dry matter when infested with B. tabaci and C. includens or by C. includens only. B. tabaci feeding did not alter contents of chlorophyll and carotenoids, and perhaps the reduction in plant growth was related to salivary toxins. Concluding, both Bt and non-Bt soybean plants were susceptible to B. tabaci feeding, evidencing necessity of developing soybean cultivars resistant to B. tabaci. Resumo A soja Bt (transgênica) foi desenvolvida para controlar insetos-praga como Anticarsia gemmatalis e Chrysodeixis includens. Este objetivo foi alcançado com sucesso; no entanto, recentemente, alguns autores afirmaram que as plantas de soja Bt têm sido mais suscetíveis do que a soja não-Bt a Bemisia tabaci MEAM1. Além disso, não se sabe se plantas de soja Bt infestadas por B. tabaci tornam-se menos resistentes às pragas-alvo. Portanto, este estudo teve como objetivo avaliar: (i) se a infestação prévia de B. tabaci pode comprometer a resistência da soja Bt e não Bt a C. includens; (ii) os efeitos de infestações de B. tabaci no crescimento de plantas de soja Bt e não Bt; e (iii) se a alimentação de B. tabaci reduz os teores de clorofila e carotenoides das plantas de soja. Plantas de soja Bt e não Bt pré-infestadas com B. tabaci não apresentaram alterações na resistência a C. includens. Plantas de soja Bt infestadas com B. tabaci apresentaram menor altura de planta do que plantas não infestadas. Diferentemente, as plantas de soja não-Bt não apresentaram redução no crescimento das plantas devido à alimentação de B. tabaci. Plantas de soja Bt sofreram redução de matéria seca apenas sob infestação dupla (B. tabaci e C. includens), enquanto plantas de soja não Bt apresentaram redução de matéria seca quando infestadas por B. tabaci e C. includens ou apenas por C. includens. A alimentação de B. tabaci não alterou os teores de clorofila e carotenoides, e talvez a redução no crescimento da planta esteja relacionada a toxinas salivares. Concluindo, tanto as plantas de soja Bt quanto as não-Bt foram suscetíveis à alimentação de B. tabaci, evidenciando a necessidade do desenvolvimento de cultivares de soja resistentes a B. tabaci.

Background The efficacy of Bt crystal proteins has been compromised due to their extensive utilization in the field. The second-generation Bt vegetative insecticidal proteins could be the best-suited alternative to combat resistance build-up due to their broad range affinity with midgut receptors of insects. Material and results The codon-optimized synthetic vegetative insecticidal proteins (Vip3Aa) gene under the control of CaMV35S promoter was transformed into a locally developed transgenic cotton variety (CKC-01) expressing cry1Ac and cry2A genes. Transformation efficiency of 1.63% was recorded. The highest Vip3Aa expression (51.98-fold) was found in MS3 transgenic cotton plant. Maximum Vip3Aa protein concentration (4.23 µg/mL) was calculated in transgenic cotton plant MS3 through ELISA. The transgenic cotton plant (MS3) showed one copy number on both chromatids in the homozygous form at chromosome 8 at the telophase stage. Almost 99% mortality of H. armigera was recorded in transgenic cotton plants expressing double crystal proteins pyramided with Vip3Aa gene as contrasted to transgenic cotton plant expressing only double crystal protein with 70% mortality. Conclusions The results obtained during this study suggest that the combination of Bt cry1Ac , cry2A , and Vip3Aa toxins is the best possible alternative approach to combat chewing insects.

Insects make up the bulk of terrestrial diversity and about half of insect species are herbivores that have direct relationships with their host plants and are the basis of the entire food chain, on which wildlife and humanity depend. Some herbivorous insect traits, such as their spatio-temporal distribution, are especially relevant in the current scenario of global changes, which are more pronounced in high elevation areas, helping to improve the effectiveness of conservation actions. Here we evaluated the influence that different spatiotemporal scales have on three free-feeding herbivorous insect guilds (fluid-feeding, leaf-chewing, and xylophagous insects) in montane forest islands immersed in a grassland-dominated matrix ( campo rupestre ). We assessed whether species turnover or nestedness was the main component determining both spatial and temporal species composition variation (β-diversity) of the herbivorous insect community. We also checked the temporal effect on herbivorous insect guilds composition between vertical strata. We sampled herbivorous insects during two summers and two winters in 14 forest islands of different sizes and shapes in a natural mountainous fragment located in southeastern Brazil. A total of 6597 herbivorous insects representing 557 morphospecies were sampled, 290 of which were fluid-feeding, 147 leaf-chewing and 120 xylophagous insects. We found a main contribution of time scale in the organization of the herbivorous insect composition sampled in this study, mainly by turnover, with small differences among guilds. Additionally, we could see that climate determined the local variation of species, corroborating that we have a highly variable always-green system over space and time where the understory community varies less in comparison to the canopy community. Our findings suggest that long-term ecological research on herbivorous community structure in relation to climatic variation is a key element for future investigations, which can be decisive for the conservation of herbivorous insect communities. We also suggest that the effects of anthropogenic pressures must be monitored in this system, since these forest islands may serve as warming refuges in a fragmented landscape holding an invaluable diversity of species that, without these old-growth forest reservoirs, would be doomed to disappear.

This study evaluated whether herbivorous insects can be expected to have particular adaptations to withstand the harsh dry season in tropical dry forests (TDFs). We specifically investigated a possible escape in space, with herbivorous insects moving to the few evergreen trees that occur in this ecosystem; and escape in time, with herbivores presenting an increased nocturnal rather than diurnal activity during the dry season. We determined the variation in the free-feeding herbivorous insects (sap-sucking and leaf chewing) between seasons (beginning and middle of both rainy and dry seasons), plant phenological groups (deciduous and evergreen trees) and diel period (diurnal and nocturnal) in a Brazilian TDF. We sampled a total of 5827 insect herbivores in 72 flight-interception traps. Contrary to our expectations, we found a greater herbivore diversity during the dry season, with low species overlap among seasons. In the dry season, evergreen trees supported greater richness and abundance of herbivores as compared to deciduous trees. Insects were also more active at night during the dry season, but no diel differences in insect abundance were detected during the rainy season. These results indicate that the strategies used by insect herbivores to withstand the severe climatic conditions of TDFs during the dry season include both small-scale escape in space and time, with evergreen trees playing a key role in maintaining resident insect herbivore populations in TDFs. Relatively more nocturnal activity during the dry season may be related to the avoidance of harsh climatic conditions during the day. We suggest that the few evergreen tree species occurring in the TDF landscape should be especially targeted for protection in this threatened ecosystem, given their importance for insect conservation.

Background The rice gall midge (RGM, Orseolia oryzae , Wood-Mason), an important stem-feeding pest worldwide, has caused serious production losses over the past decades. Rice production practices indicate that the most reliable method for managing RGM is the deployment of cultivars that incorporate host resistance. However, the conventional phenotypic screening method of rice resistance to RGM suggested by the International Rice Research Institute (IRRI) has been used for approximately 30 years, and only 12 rice varieties/lines (including controls) can be evaluated in one tray. It is not suitable for high-throughput phenotyping of rice germplasm. Moreover, a suitable method to prepare samples for molecular biological studies of rice resistance against RGM is imperative with the rapid development of modern molecular techniques. Results The proper density of seedlings/RGM was determined for four seeding arrangements. A high-throughput phenotyping method (HTPM) for 60 lines/varieties infested with 36 female RGM adults in one tray, as described by method 4–3 (seeded 60 lines/varieties), was developed and verified using mutant screening. Furthermore, one RGM resistance gene flanked by markers 12RM28346 and 12RM28739 on chromosome 12 was simultaneously detected using method 2–2 (seeded 30 lines/varieties in one tray) treated with 24 RGM and analyzed using conventional and simplified grading systems. Genetic analysis of the RGM resistance gene was confirmed using a method identical to that suggested by IRRI. Finally, one bucket with 24 seedlings treated with at least five female RGM adults was efficacious and could offer adequate samples for insect development observation or molecular biological studies. Conclusion A highly efficient and reliable procedure for evaluation of resistance in rice to RGM was developed and improved, and was verified through mutant screening, gene mapping, genetic analysis, and insect growth and development observations.

Caterpillar feeding induces direct and indirect defences in brassicaceous plants. This study focused on the role of the octadecanoid pathway in induced indirect defence in Brassica oleracea. The effect of induction by exogenous application of jasmonic acid (JA) on the responses of Brussels sprouts plants and on host-location behaviour of associated parasitoid wasps was studied. Feeding by the biting-chewing herbivores Pieris rapae and Plutella xylostella resulted in significantly increased endogenous levels of JA, a central component in the octadecanoid signalling pathway that mediates induced plant defence. The levels of the intermediate 12-oxophyto-dienoic acid (OPDA) were significantly induced only after P. rapae feeding. Three species of parasitoid wasps, Cotesia glomerata, C. rubecula, and Diadegma semiclausum, differing in host range and host specificity, were tested for their behavioural responses to volatiles from herbivore-induced, JA-induced, and non-induced plants. All three species were attracted to volatiles from JA-induced plants compared with control plants; however, they preferred volatiles from herbivore-induced plants over volatiles from JA-induced plants. Attraction of C. glomerata depended on both timing and dose of JA application. JA-induced plants produced larger quantities of volatiles than herbivore-induced and control plants, indicating that not only quantity, but also quality of the volatile blend is important in the host-location behaviour of the wasps.

In the search for environmentally friendly options to manage plant pests and enhance productivity with fewer external inputs in agriculture, the screening of landraces has gained importance. The aims of this study were to characterize functional leaf traits in maize landraces, including morphological traits such as leaf toughness, thickness, specific leaf area (SLA), and number of trichomes, as well as physiological leaf traits such as photosynthesis, transpiration rate, and stomatal conductance. We aimed to determine the association of these traits with leaf damage caused by Spodoptera frugiperda across 24 maize landraces. Relationships of dependence among leaf traits and leaf damage were determined using principal component analysis, multiple regression analysis and sequential path analyses. All morphological leaf traits showed differences among maize landraces mainly at 20 and 30 days after emerge (DAE) whereas at 40 DAE the differences decreased. By the contrast, physiological leaf traits had high variation at 40 DAE among maize landraces. We found a negative dependency of principal component conformed by photosynthesis, transpiration rate, and stomatal conductance over leaf damage. Based on the level of leaf damage, we categorized maize landraces into two groups with low and high susceptibility. Surprisingly, we observed that in the former group, physiological leaf traits affected the number of trichomes, whereas in the latter group, they influenced leaf thickness. This finding highlights the necessity of detailed assessments of foliage properties to understand the impact of S. frugiperda infestations. Such an analytical approach is crucial in scientific research, providing deeper insights into the underlying mechanisms of interaction between the pest and its host plants.

Plant responses to herbivores are complex. Genes activated on herbivore attack are strongly correlated with the mode of herbivore feeding and the degree of tissue damage at the feeding site. Phloem-feeding whiteflies and aphids that produce little injury to plant foliage are perceived as pathogens and activate the salicylic acid (SA)-dependent and jasmonic acid (JA)/ethylene-dependent signaling pathways. Differential expression of plant genes in response to closely related insect species suggest that some elicitors generated by phloem-feeding insects are species-specific and are dependent on the herbivore's developmental stage. Other elicitors for defense-gene activation are likely to be more ubiquitous. Analogies to the pathogen-incompatible reactions are found. Chewing insects such as caterpillars and beetles and cell-content feeders such as mites and thrips cause more extensive tissue damage and activate wound-signaling pathways. Herbivore feeding is not equivalent to mechanical wounding. Wound responses are a part of the induced responses that accompany herbivore feeding. Herbivores induce direct defenses that interfere with herbivore feeding, growth and development, fecundity, and fertility. In addition, herbivores induce an array of volatiles that creates an indirect mechanism of defense. Volatile blends provide specific cues to attract herbivore parasites and predators to infested plants. The nature of the elicitors for volatile production is discussed.