Insect Diversity Research Articles

A new suite of reporter vectors and a novel landing site survey system to study cis-regulatory elements in diverse insect species

Comparative analyses between traditional model organisms, such as the fruit fly Drosophila melanogaster, and more recent model organisms, such as the red flour beetle Tribolium castaneum, have provided a wealth of insight into conserved and diverged aspects of gene regulation. While the study of trans-regulatory components is relatively straightforward, the study of cis-regulatory elements (CREs, or enhancers) remains challenging outside of Drosophila. A central component of this challenge has been finding a core promoter suitable for enhancer-reporter assays in diverse insect species. Previously, we demonstrated that a Drosophila Synthetic Core Promoter (DSCP) functions in a cross-species manner in Drosophila and Tribolium. Given the over 300 million years of divergence between the Diptera and Coleoptera, we reasoned that DSCP-based reporter constructs will be useful when studying cis-regulation in a variety of insect models across the holometabola and possibly beyond. To this end, we sought to create a suite of new DSCP-based reporter vectors, leveraging dual compatibility with piggyBac and PhiC31-integration, the 3xP3 universal eye marker, GATEWAY cloning, different colors of reporters and markers, as well as Gal4-UAS binary expression. While all constructs functioned properly with a Tc-nub enhancer in Drosophila, complications arose with tissue-specific Gal4-UAS binary expression in Tribolium. Nevertheless, the functionality of these constructs across multiple holometabolous orders suggests a high potential compatibility with a variety of other insects. In addition, we present the piggyLANDR (piggyBac-LoxP AttP Neutralizable Destination Reporter) platform for the establishment of proper PhiC31 landing sites free from position effects. As a proof-of-principle, we demonstrated the workflow for piggyLANDR in Drosophila. The potential utility of these tools ranges from molecular biology research to pest and disease-vector management, and will help advance the study of gene regulation beyond traditional insect models.

Establishment of RNAi-Mediated Pest Control Method for Red Imported Fire Ant, Solenopsis invicta.

RNAi plays a crucial role in insect gene function research and pest control field. Nonetheless, the variable efficiency of RNAi across diverse insects and off-target effects also limited its further application. In this study, we cloned six essential housekeeping genes from Solenopsis invicta and conducted RNAi experiments by orally administering dsRNA. Then, we found that mixing with liposomes significantly enhanced the RNAi efficiency by targeting for SiV-ATPaseE. Additionally, we observed a certain lethal effect of this dsRNA on queens by our established RNAi system. Furthermore, no strict sequence-related off-target effects were detected. Finally, the RNAi effect of large-scale bacteria expressing dsRNA was successfully confirmed for controlling S. invicta. In summary, this study established an RNAi system for S. invicta and provided a research template for the future development of nucleic acid drugs based on RNAi.

The effect of local habitat and spatial connectivity on urban seed predation.

During the last centuries, the area covered by urban landscapes is increasing all over the world. Urbanization can change local habitats and decrease connectivity among these habitats, with important consequences for species interactions. While several studies have found a major imprint of urbanization on plant-insect interactions, the effects of urbanization on seed predation remain largely unexplored. We investigated the relative impact of sunlight exposure, leaf litter, and spatial connectivity on predation by moth and weevil larvae on acorns of the pedunculate oak across an urban landscape during 2018 and 2020. We also examined whether infestations by moths and weevils were independent of each other. While seed predation varied strongly among trees, seed predation was not related to differences in sunlight exposure, leaf litter, or spatial connectivity. Seed predation by moths and weevils was negatively correlated at the level of individual acorns in 2018, but positively correlated at the acorn and the tree level in 2020. Our study sets the baseline expectation that urban seed predators are unaffected by differences in sunlight exposure, leaf litter, and spatial connectivity. Overall, our findings suggest that the impact of local and spatial factors on insects within an urban context may depend on the species guild. Understanding the impact of local and spatial factors on biodiversity, food web structure, and ecosystem functioning can provide valuable insights for urban planning and management strategies aimed at promoting urban insect diversity.

Assessing the spatial knowledge gaps of Odonata diversity and conservation in the South American Pampa

Abstract Anthropogenic activities highly threaten neotropical freshwater habitats. Aquatic insects are sensitive to environmental alterations and thus considered under great risk due to such pressures. However, large‐scale gaps regarding the knowledge of the distribution of aquatic insect species remain in the Neotropics, particularly in the South American temperate grasslands (Pampas), hindering the knowledge of hotspots of aquatic insect diversity and the prioritization of conservation areas destined to protect their populations. Here, we review the current knowledge regarding the geographical distribution of Odonata species and use ecological niche models to estimate the areas with the highest potential richness of Odonata species in the Pampas. We also calculate a sampling effort index and interpolate it with maps of protected areas to identify the spatial knowledge gaps regarding their conservation in the region Our compilation of Odonata species records retrieved 237 species for the Pampa. However, Odonata species were recorded in less than 2% of the Pampa. Our findings indicated a higher potential diversity of Odonata in northeastern Argentina, southern Uruguay and southern Brazil. Interpolations of sampling effort index and protected areas showed that 95% of the area potentially rich in Odonata species fall outside protected areas. Our study highlights the shortfall regarding the knowledge of the distribution of Odonata diversity in non‐forest landscapes of the Neotropical region. It also shows that protected areas of the Pampa cover a minor area of the Odonata species range in the region.

Peran Serangga Tanah dalam Budidaya Talas Beneng di Karangtanjung

Araceae is a plant family known as taro with a high level of diversity and is easily found by the public. Beneng taro (Xanthosoma undipes) is a plant in the Araceae family that is included in the local food commodities originating from Banten Province, especially in the Mount Karang. Soil insects are a type of insects that have an essential role in the food cycle, which supports the weathering process of organic material to have a positive influence on the physical and chemical properties of soil. The diversity of soil insects on taro beneng plants in Cinyurup Village is essential to be known to open up new opportunities for understanding their ecological benefits related to agricultural sustainability and biodiversity conservation. This research aims to determine soil insects and their role in the beneng taro cultivation area and to determine the diversity index of soil insects in the area. This research was carried out in February-March in Cinyurup Village, Karang Tanjung District, Pandeglang Regency. This research includes quantitative descriptive research with an exploratory method. Sampling was carried out by purposive sampling using the pitfall trap technique. The samples in this research are all ground insects trapped in pitfall traps. Trapped samples were identified based on their role and the calculated diversity index of the Shannon-Wiener method, which can facilitate the analysis of information about the type of individual and the number in an area. The identification results showed that there were 252 insects belonging to 6 families, namely Formicidae, Gryllidae, Tetrigidae, Scarabaeidae, Anisolabidae, and Muscidae. Formicidae and Scarabaeidae are land insects found in abundance due to the farming system based on the cultivation of wire that supports their survival. The diversity index value was obtained at 1.03064 which is included in the medium category. The insects found have varied roles, namely predators (Formicidae, Anisolabidae), herbivores (Gryllidae, Tetrigidae), pollinators (Formicidae, Muscidae), and decomposers (Scarabaeidae).

Different Sensitivity of Flower-Visiting Diptera to a Neonicotinoid Insecticide: Expanding the Base for a Multiple-Species Risk Assessment Approach.

Insects play an essential role as pollinators of wild flowers and crops. At the same time, pollinators in agricultural environments are commonly exposed to pesticides, compromising their survival and the provision of pollination services. Although pollinators include a wide range of species from several insect orders, information on pesticide sensitivity is mostly restricted to bees. In addition, the disparity of methodological procedures used for different insect groups hinders the comparison of toxicity data between bees and other pollinators. Dipterans are a highly diverse insect order that includes some important pollinators. Therefore, in this study, we assessed the sensitivity of two hoverflies (Sphaerophoria rueppellii, Eristalinus aeneus) and one tachinid fly (Exorista larvarum) to a neonicotinoid insecticide (Confidor®, imidacloprid) following a comparative approach. We adapted the standardized methodology of acute contact exposure in honey bees to build dose-response curves and calculate median lethal doses (LD50) for the three species. The methodology consisted in applying 1 µL of the test solution on the thorax of each insect. Sphaerophoria rueppelli was the most sensitive species (LD50 = 10.23 ng/insect), and E. aeneus (LD50 = 18,176 ng/insect) the least. We then compared our results with those available in the literature for other pollinator species using species sensitivity distribution (SSD). Based on the SSD curve, the 95th percentile of pollinator species would be protected by a safety factor of 100 times the Apis mellifera endpoint. Overall, dipterans were less sensitive to imidacloprid than most bee species. As opposed to most bee species, oviposition and fecundity of many dipteran species can be reliably assessed in the laboratory. We measured the number of eggs laid following exposure to different insecticide doses and assessed the potential trade-off between oviposition and survival through the sublethal sensitivity index (SSI). Exposure to imidacloprid had a significant effect on fecundity, and SSI values indicated that oviposition is a sensitive endpoint for the three dipteran species tested. Future studies should integrate this information related to population dynamics in simulation models for environmental risk assessment.

Polymorphic Clustering and Approximate Masking Framework for Fine-Grained Insect Image Classification

Insect diversity monitoring is crucial for biological pest control in agriculture and forestry. Modern monitoring of insect species relies heavily on fine-grained image classification models. Fine-grained image classification faces challenges such as small inter-class differences and large intra-class variances, which are even more pronounced in insect scenes where insect species often exhibit significant morphological differences across multiple life stages. To address these challenges, we introduce segmentation and clustering operations into the image classification task and design a novel network model training framework for fine-grained classification of insect images using multi-modality clustering and approximate mask methods, named PCAM-Frame. In the first stage of the framework, we adopt the Polymorphic Clustering Module, and segmentation and clustering operations are employed to distinguish various morphologies of insects at different life stages, allowing the model to differentiate between samples at different life stages during training. The second stage consists of a feature extraction network, called Basenet, which can be any mainstream network that performs well in fine-grained image classification tasks, aiming to provide pre-classification confidence for the next stage. In the third stage, we apply the Approximate Masking Module to mask the common attention regions of the most likely classes and continuously adjust the convergence direction of the model during training using a Deviation Loss function. We apply PCAM-Frame with multiple classification networks as the Basenet in the second stage and conduct extensive experiments on the Insecta dataset of iNaturalist 2017 and IP102 dataset, achieving improvements of 2.2% and 1.4%, respectively. Generalization experiments on other fine-grained image classification datasets such as CUB200-2011 and Stanford Dogs also demonstrate positive effects. These experiments validate the pertinence and effectiveness of our framework PCAM-Frame in fine-grained image classification tasks under complex conditions, particularly in insect scenes.

Studies on Neotropical Phasmatodea XXVI: Taxonomic review of Cladomorformia tax. n., a lineage of Diapheromerinae stick insects, with the descriptions of seven new genera and 41 new species (Phasmatodea: Occidophasmata: Diapheromerinae)

Studies on Neotropical Phasmatodea XXVI: Taxonomic review of Cladomorformia tax. n., a lineage of Diapheromerinae stick insects, with the descriptions of seven new genera and 41 new species (Phasmatodea: Occidophasmata: Diapheromerinae)

Characterisation of a Novel Insect-Specific Virus Discovered in Rice Thrips, Haplothrips aculeatus.

Insects constitute the largest proportion of animals on Earth and act as significant reservoirs and vectors in disease transmission. Rice thrips (Haplothrips aculeatus, family Phlaeothripidae) are one of the most common pests in agriculture. In this study, the full genome sequence of a novel Ollusvirus, provisionally named "Rice thrips ollusvirus 1" (RTOV1), was elucidated using transcriptome sequencing and the rapid amplification of cDNA ends (RACE). A homology search and phylogenetic tree analysis revealed that the newly identified virus is a member of the family Aliusviridae (order Jingchuvirales). The genome of RTOV1 contains four predicted open reading frames (ORFs), including a polymerase protein (L, 7590 nt), a glycoprotein (G, 4206 nt), a nucleocapsid protein (N, 2415 nt) and a small protein of unknown function (291 nt). All of the ORFs are encoded by the complementary genome, suggesting that the virus is a negative-stranded RNA virus. Phylogenetic analysis using polymerase sequences suggested that RTOV1 was closely related to ollusvirus 1. Deep small RNA sequencing analysis reveals a significant accumulation of small RNAs derived from RTOV1, indicating that the virus replicated in the insect. According to our understanding, this is the first report of an Ollusvirus identified in a member of the insect family Phlaeothripidae. The characterisation and discovery of RTOV1 is a significant contribution to the understanding of Ollusvirus diversity in insects.

Identification and Genetic Diversity of Spodoptera frugiperda J. E. Smith (Lepidoptera: Noctuidae) in Egypt

Fall armyworm, Spodoptera frugiperda, is a serious agricultural pest native to tropical and subtropical regions of the Western Hemisphere and has invaded Africa and further spread into most countries of Asia within two years. In Egypt, researchers have conducted thorough investigations into the behavior of the fall armyworm and various methods to manage its impact. This study aims to further our understanding of the genetic diversity of fall armyworm populations in Egypt. By collecting specimens from five provinces across the country, we sought to analyze their genetic makeup. Through examination of partial sequences of the mitochondrial cytochrome oxidase subunit I (COI), we identified three distinct haplotypes. Phylogenetic analysis suggests that the primary sources of Spodoptera frugiperda in Egypt likely stem from both Asian and African origins. Employing the PCR-RFLP technique on the complete COI sequence, we were able to discern genotype patterns within the fall armyworm population. Our findings indicate the presence of two distinct strains—the Corn and Rice strains—within Egypt. This research contributes essential insights into the genetic diversity of insects in Egypt, providing valuable knowledge that can inform more effective strategies for managing this agricultural pest.

Innovative Exploration of a Bio-Inspired Sensor Fusion Algorithm: Enhancing Micro Satellite Functionality through Touretsky's Decentralized Neural Networks

Insect-inspired sensor fusion algorithms have presented a promising avenue in the development of robust and efficient systems, owing to the insects' ability to process numerous streams of noisy sensory data. The ring attractor neural network architecture has been identified as a noteworthy model for the optimal integration of diverse insect sensors. Expanding on this, our research presents an innovative bio-inspired ring attractor neural network architecture designed to augment the performance of microsatellite attitude determination systems through the fusion of data from multiple gyroscopic sensors.Extensive simulations using a nonlinear model of the microsatellite, while incorporating specific navigational disturbances, have been conducted to ascertain the viability and effectiveness of this approach. The results obtained have been superior to those of alternative methodologies, thus highlighting the potential of our proposed bio-inspired fusion technique. The findings indicate that this approach could significantly improve the accuracy and robustness of microsatellite systems across a wide range of applications.

Hedgerow Gardens Provide Floral Resources for Diverse Insect Visitors to Avocado Fowers in Southern California

Hedgerow Gardens Provide Floral Resources for Diverse Insect Visitors to Avocado Fowers in Southern California

Hedgerow Gardens Provide Floral Resources for Diverse Insect Visitors to Avocado Flowers in Southern California

Hedgerow Gardens Provide Floral Resources for Diverse Insect Visitors to Avocado Flowers in Southern California

Molecular identification of Hymenopteran insects collected by using Malaise traps from Hazarganji Chiltan National Park Quetta, Pakistan.

The order Hymenoptera holds great significance for humans, particularly in tropical and subtropical regions, due to its role as a pollinator of wild and cultivated flowering plants, parasites of destructive insects and honey producers. Despite this importance, limited attention has been given to the genetic diversity and molecular identification of Hymenopteran insects in most protected areas. This study provides insights into the first DNA barcode of Hymenopteran insects collected from Hazarganji Chiltan National Park (HCNP) and contributes to the global reference library of DNA barcodes. A total of 784 insect specimens were collected using Malaise traps, out of which 538 (68.62%) specimens were morphologically identified as Hymenopteran insects. The highest abundance of species of Hymenoptera (133/538, 24.72%) was observed during August and least in November (16/538, 2.97%). Genomic DNA extraction was performed individually from 90/538 (16.73%) morphologically identified specimens using the standard phenol-chloroform method, which were subjected separately to the PCR for their molecular confirmation via the amplification of cytochrome c oxidase subunit 1 (cox1) gene. The BLAST analyses of obtained sequences showed 91.64% to 100% identities with related sequences and clustered phylogenetically with their corresponding sequences that were reported from Australia, Bulgaria, Canada, Finland, Germany, India, Israel, and Pakistan. Additionally, total of 13 barcode index numbers (BINs) were assigned by Barcode of Life Data Systems (BOLD), out of which 12 were un-unique and one was unique (BOLD: AEU1239) which was assigned for Anthidium punctatum. This indicates the potential geographical variation of Hymenopteran population in HCNP. Further comprehensive studies are needed to molecularly confirm the existing insect species in HCNP and evaluate their impacts on the environment, both as beneficial (for example, pollination, honey producers and natural enemies) and detrimental (for example, venomous stings, crop damage, and pathogens transmission).

Succession and seasonality drive tropical butterfly assembly after an extreme hurricane

AbstractThe composition and biodiversity of insect community assemblages are mediated by a complex set of biotic and abiotic factors. Among these factors are forest structure and atmospheric variables (like temperature and humidity), which are heavily influenced by frequent hurricane activity in the Caribbean. Despite this, changes in Caribbean insect assemblages as forests recover from hurricane disturbance are poorly documented. Butterflies represent a charismatic model taxon in biodiversity and conservation, and are thus an ideal subject for exemplifying these shifts in insect abundances and diversity across ecological succession. Here, we monitored butterfly communities in two Puerto Rican forests differing in structure (i.e., canopy height, tree size) to assess butterfly diversity, abundances, and community level wing traits (size and color) over 1 year, beginning 6 months after Hurricane Maria. Monthly sampling over the course of 1 year revealed no relationships between abundances and canopy openness or humidity; instead, species abundances fluctuated seasonally and were nonlinearly correlated with temperature. In contrast, wing size and color were linearly correlated with abiotic variables. Specifically, wings were larger in cooler and more open conditions. Wing color saturation and brightness were negatively correlated with humidity. Our results suggest that, first, a functional approach may provide better insight into the factors mediating species responses to disturbances. Second, further disentangling abundance seasonality from impacts of extreme disturbances necessitates long‐term monitoring.Abstract in Spanish is available with online material.

Trends in the Brazilians' engagement with insects on the internet

Abstract It is estimated that the tropical and south temperate regions harbour approximately 85% of the total entomofauna, and Brazil, which is the largest country in the Neotropical region, shows the greatest insect diversity in the world. However, to the best of our knowledge, the Brazilian population's interest in insects is yet to be investigated. We use Google Trends platform for prospecting temporal variation in the public engagement with nature. From searches in this database, from 2012 to 2020, using common names of 21 insect groups, we evaluated: (I) Brazilians' internet interest on insects over time, (II) the highest search themes for each insect group and (III) which pairs of insects are positively or negatively correlated regarding the public interest through years. Our findings demonstrate that Brazilians' internet interest regarding the studied insect groups is constant over time, which may be linked with the low attention these organisms generally receive, and that Brazilians are prone to perceive them as harmful, as we can observe in many of the top queries related to studied groups. We observed a considerable synchronous growth in interest regarding pairs of insects considered as domestic pests, medically important and charismatic, and the searches on these groups referred mainly to their biological, ecological and cultural aspects. Our results indicate that investment in environmental education in Brazil can motivate public engagement with insects. Also, our data can be crucial for directing conservation policies for insects in the tropical region.

Diversity pattern of insects from Macao based on an updated species checklist after 25 years.

Insects represent one of the most diverse groups in the organism world with extremely rich species and morphological diversity, playing important roles in natural and city ecosystems. Regional compilation of insect species lists helps to clarify the richness of insect species in a region, enhances our understanding the structure and function of a local ecosystem and promotes the protection and development of insect resources. Moreover, it also serves as a valuable reference for cities with small area, large population and high urbanisation like Macao. Macao (Macau) Special Administrative Region (SAR) is situated at the Pearl River Delta on the southeast coast of mainland China. With urban development accelerating at great rate in a quite restricted area, Macao still has rich fauna, within which the insect diversity is surprisingly high. In this study, we systematically sorted out major references items of manuals or handbooks, monographs, articles, dissertations, official websites and other publicly available information sources about the insects recorded in Macao and, thus, generated a checklist of 15 orders, 166 families, 868 genera, 1,339 species and 118 subspecies. During this process, the preliminarily summarised list was re-examined to eliminate synonyms and invalid species, based on many more extensive literature reviews. Besides, spelling errors of scientific names, authors and years were corrected. Meanwhile, the catalogue revealed a different composition pattern of species diversity between orders from those of the world and China. Even based on the most conservative estimates, the number of insect species in Macao should not be lower than 3,340 species, which hints at the necessity of deeper investigations with adequate collecting in the future to achieve more comprehensive recognition and understanding of Macao's insect biodiversity.

Entomofauna Associated with Corn Cultivation and Damage Caused by Some Pests According to the Planting Season on the Ecuadorian Coast

Corn (Zea mays L.) is an important cereal for food sovereignty, extensively planted due to its adaptation to various agroecological conditions. Climatic conditions and pests can affect its production. Concerning the latter, natural enemies could be considered in biological control programs. The objective of this study was to estimate the influence of the presence of insects, damage to plants and cobs, and their incidence on corn yield in two planting seasons. During the rainy (February to May) and dry seasons (Jun to October) of the year 2023, in Calceta, Lodana, and Quevedo, three important corn-growing areas on the Ecuadorian coast, corn plots of 1126 m2 were planted. The diversity of phytophagous and natural enemies was estimated. Damage to young plants and cobs was observed, and corn yield was determined. Of the 9073 insects observed, 44.2% and 55.8% constituted phytophagous and natural enemies, respectively. A moderate diversity (Shannon-H Index = 2.474–2.629 and Margalef Index = 2.734–3.110) of insects associated with corn was determined. Yield (range: 6.9 to 15.3 t) was negatively correlated with rainfall and cob damage (p < 0.05). Although precipitation is necessary for planting in rainy season, frequent and intense rains could be affect corn yield.

Entomocomplex of Radish in Open-field Cultivation in the Central Part of the Right-Bank Forest-Steppe of Ukraine

Goal. To specify the species composition of insect pests affecting radish (Raphanus sativus (L.) convar. radicula (Pers) Sazon.) in open-field cultivation conditions in the central part of the Right-Bank Forest-Steppe of Ukraine.
 Methods. The study was conducted following commonly accepted methods in entomology and vegetable growing. From 2008 to 2022, we investigated the species diversity of insects, dominant species, their abundance, and the structure of the harmful entomocomplex in radish crops grown in open-field conditions on household plots and farms in the Cherkasy region.
 Results. In the entomological complex of radish, 59 species of phytophagous insects from 20 families and 8 orders were identified: Coleoptera — 29%, Lepidoptera — 27%, Homoptera — 15%, Diptera — 10%, Orthoptera — 9%, Thysanoptera — 5%, Hemiptera — 3%, Hymenoptera — 2%. Sixteen dominant species of phytophagous insects were found in radish crops, including diamondback moth (Plutella maculipennis Curt.), crucifer flea beetles (Phyllotreta cruciferae Goeze), striped flea beetle (Phyllotreta undulata Kutsch.), cabbage white butterfly (Pieris brassicae L.), turnip moth (Agrotis segetum Denis&Schiff.) and owlet moth (Lacanobia oleracea L.), crucifer shield bug (Eurydema ventralis Kol.), cabbage aphid (Brevicoryne brassicae L.), spring root maggot (Delia brassicae Bouche) and summer root maggot (Delia floralis Fallen), seedcorn maggot (Delia platura Mg.), sawfly (Athalia rosae L.), stem weevil (Ceutorrhynchus quadridens Panz.), tobacco thrips (Thrips tabaci Lindeman), turnip sawfly (Entomoscelis adonidis Pallas), and crucifer webworm (Evergestis extimalis Scop.). Through multi-year studies, the periods of insect damage corresponding to radish developmental stages were determined. The most critical damage to radish plants by harmful insects occurred during the initial stages of development (ВВСН 0—9) and from emergence to the growth and formation of the root crop (BBCH 42—48).
 Conclusions. It was found that among the identified phytophagous insects in radish crops, the most economically significant damage was caused by the cabbage aphid, crucifer flea beetles, striped flea beetle, spring cabbage fly, and seedcorn maggot. On average over the research period (2008—2022), the largest areas of radish crops were infested by the diamondback moth, crucifer flea beetles, and striped flea beetle, cabbage white butterfly, winter and owlet moths, cabbage shield bug, and cabbage aphid, ranging from 5% to 80%, reaching a maximum in some years up to 60—100% of the area.

New descriptions of the larval and pupal stages of Orthocladiusnitidoscutellatus and Psectrocladiusnevalis from Xizang, China (Diptera, Chironomidae).

Tibetan Plateau is one of the most typical areas of biodiversity in the world because of its unique environmental and regional units, which breed unique biological communities and concentrate on many unique and rare wild animals and plants. Research on Chironomidae in the Tibetan Plateau is relatively weak. At present, the identification of Chironomidae species mainly depends on male adults, while identification of larvae and pupae is relatively difficult and there is less research on them. During the investigations of insect diversity in the Tibetan Plateau, larval and pupal stages of Orthocladiusnitidoscutellatus Lundström, 1915 and Psectrocladiusnevalis Akhrorov, 1977 were described and illustrated. Matching and identification of larval and pupal stages were based on DNA barcodes. Neighbour-joining trees were reconstructed, based on known Orthocladius and Psectrocladius COI DNA barcodes, respectively.